Fatty acids, tall-oil, magnesium salts

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Salmonella Typhimurium Reverse Mutation Test

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2005-08-22 - 2005-09-15

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Justification for read-across, see attached file.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

21 july, 1997

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Tall oil

EC Number:

232-304-6

EC Name:

Tall oil

Cas Number:

8002-26-4

Molecular formula:

Not applicable - UVCB substance

Test material form:

liquid: viscous

Method

Species / strain

Species / strain / cell type:

other: Salmonella typhimurium TA97a, TA98, TA100, TA102 and TA153

Metabolic activation:

with and without

Metabolic activation system:

S9-mix

Test concentrations with justification for top dose:

Test substance (all strains: 1st experiment; TA98, TA100, TA102 and TA1535: 2nd experiment)

Dose: 5000, 1667, 556, 185 and 62 µg/plate

Number of samples: 3 samples

Test substance (TA97a: 2nd experiment)

Dose: 556, 185, 62, 21, 7 and 2.3 µg/plate

Number of samples: 3 samples

Justification: The concentrations for the first experiment were set according to a preliminary toxicity test, see 3.2.
The test substance was not toxic up to 5000 µg/petri dish.

It was therefore decided to use 5000 µg/plate as the highest concentration which is the limit concentration according to the guidelines. Each of the other 4 concentrations was 1/3 of the preceding one.

For the second experiment the concentrations were changed only for strain TA97a according to the first experiment. The test substance was toxic to strain TA97a at 5000 and 1667 µg/plate. Thus it was decided to use 556 µg/plate as the highest concentration for the second experiment, which could be in the toxic range and to use a total of 6 concentrations.

The number and intervals of the concentrations are in accordance with the guidelines (at least 5 concentrations with intervals of about 10 ).

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

other: 4-Nitro-o-phenylene-diamine, t-Butyl-hydroperoxide, 2- Aminoanthracene, 1,8-Dihydroxy-anthraquinone, 7,12-Dimethylbenz[a]anthracene, 2-Nitrofluorene

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 97

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:

S. typhimurium TA 98

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 3

Species / strain:

S. typhimurium 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 4

Species / strain:

S. typhimurium TA 102

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 5

Species / strain:

S. typhimurium TA 1535

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:

In the preliminary test with strain TA100 no toxicity was seen up to 5000 µg/plate.

In the main test the test substance was only toxic to strain TA97a, resulting in a completely or almost missing bacterial background lawn at 5000 and 1667 µg/plate. At 556 µg/plate and beneath the bacterial background was normal.

Applicant's summary and conclusion

Conclusions:

According to these results, "CRUDE TALL OIL" is not mutagenic in the Ames test with the strains of Salmonella typhimurium TA97a, TA98, TA100, TA102 and TA1535 with and without an external metabolising system up to 5000 µg/plate.

Executive summary:

Method

"CRUDE TALL OIL" was tested for mutagenic activity with the "Salmonella typhimurium Reverse Mutation Test" (Ames Test). The study was conducted in accordance with the OECD-guideline 471 and directive 2000/32/EC, part B.13/14.

The test substance was dissolved in DMSO. The following concentrations were tested:

62, 185, 556, 1667 and 5000 µg per plate without external metabolisation, and

62, 185, 556, 1667 and 5000 µg per plate with S9-mix from Aroclor 1254 induced microsomes of rat liver as an external metabolising system.

In the second experiment the concentrations were changed for strain TA97a as follows:  

2.3, 7, 21, 62, 185 and 556 µg per plate without external metabolisation, and

2.3, 7, 21, 62, 185 and 556 µg per plate with S9-mix from Aroclor 1254 induced microsomes of rat liver as an external metabolising system.

The test was performed according to the "direct plate incorporation method". As test system the bacterial strains Salmonella typhimurium TA97a, TA98, TA100, TA102 and TA1535 were used. Negative and positive controls were included. An independent repetition of the experiment was performed.  

Results

Positive controls:

All positive control groups showed significantly increased mutation frequencies which demonstrate the sensitivity of the test system.  

Test substance:

Toxicity:

The test substance was toxic only to strain TA97a, resulting in a completely or almost missing bacterial background lawn at 5000 and 1667 µg/plate. At 556 µg/plate and beneath the bacterial background was normal. No toxicity was noted in the other strains.

Solubility:

At the 5000, 1667 and 556 µg/plate samples a precipitate was visible when the test substance was mixed with the agar. At 5000 µg/plate the precipitate was still visible when the colonies were counted but did not impede the counting.

Mutagenicity:

In none of the concentrations tested and with none of the strains used an increase of the mutation frequency to more than the threshold values (250 % of the controls for strains TA98 and TA1535, 167 % of the controls for strains TA97a, TA100 and TA102) was obtained. Metabolic activation did not change these results.  

Conclusion

According to the results obtained in this study, "CRUDE TALL OIL" is non-mutagenic in the Ames test with the strains TA97a, TA98, TA100, TA102 and TA1535 with and without an external metabolising system up to 5000 µg/plate.