Poly(oxy-1,2-ethanediyl), α-[2-(dide- cylmethylammonio)ethyl]- .omega.- hydroxy-, propanoate (salt) (Bardap 26)

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:

April-May 1986

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1986

Report date:

1986

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

Not applicable.

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

bis(decyl)(2-hydroxyethyl)methylazanium propanoate

EC Number:

619-057-3

Cas Number:

94667-33-1

Molecular formula:

C29 H62 N O4 . C3 H5 O2

IUPAC Name:

bis(decyl)(2-hydroxyethyl)methylazanium propanoate

Constituent 2

Reference substance name:

alpha.-[2-(Didecylmethylammonio)ethyl]-.omega.-hydroxy-poly(oxy-1,2-ethanediyl) propionate

IUPAC Name:

alpha.-[2-(Didecylmethylammonio)ethyl]-.omega.-hydroxy-poly(oxy-1,2-ethanediyl) propionate

Constituent 3

Reference substance name:

N,N-Didecyl-N-methyl-poly(oxyethyl)ammonium Propionate

IUPAC Name:

N,N-Didecyl-N-methyl-poly(oxyethyl)ammonium Propionate

Constituent 4

Reference substance name:

Bardap 26

IUPAC Name:

Bardap 26

Details on test material:

The test substance, Hoe S3519 (N,N-Didecyl-N-methyl-poly(oxyethyl)ammonium propionate in aqueous/alcohol solution), was described as a clear, yellow liquid and stable at room temperature. The test substance was stored in the dark at 22°C.

Method

Target gene:

Reversion to histidine dependence

Species / strain

Species / strain 1

Species / strain / cell type:

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

Additional strain / cell type characteristics:

not applicable

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1538

Additional strain / cell type characteristics:

not applicable

Species / strain 3

Species / strain / cell type:

E. coli WP2 uvr A

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

S9 mix (Aroclor 1254 induced Sprague-Dawley rats)

Test concentrations with justification for top dose:

Cytotoxicity testing: 0, 4, 20, 100, 500, 2500, 10000 µg/plate
Experiment 1: 0, 4, 20, 100, 500, 2500, 10000 µg/plate
Experiment 2: 0, 0.032, 0.16, 0.8, 4, 20, 100 µg/plate

Vehicle / solvent:

Double distilled water

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

other: 2-aminothracene. With S9: TA1538, TA98, TA100, TA1537, WP2uvrA

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Remarks:

Migrated to IUCLID6: With S9: TA1535, TA1537, TA1538, TA98, TA100, WP2uvrA

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

Migrated to IUCLID6: Without S9: TA1538, TA98

Controls 4

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

Migrated to IUCLID6: Without S9: TA1537

Controls 5

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

Migrated to IUCLID6: Without S9: TA1535, TA100

Controls 6

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: MNNG. Without S9: WP2uvrA

Details on test system and experimental conditions:

Cytotoxicity was evaluated in a preliminary study with and without S9, the parameters assessed were thinning of background lawn and reduction in colony numbers. Following this, two independent assays were performed both with and without S9.

Bacteria were grown overnight in nutrient broth at 37°C. The suitable amount of bacteria in the cell suspension was checked by nephelometry. For inoculation, stock cultures stored at -80°C were used.

Top agar was prepared for the Salmonella strains by mixing 100 ml agar with 10 ml of a 0.5 mM histidine-biotin solution. With E. coli histidine was replaced with tryptophan (2.5 ml, 0.5 mM). The following ingredients were added to 2 ml of molton top agar at 45°C: 0.1 ml of overnight broth culture of tester strain, 0.1 ml test compound solution, 0.5 ml S9 mix or buffer. After mixing, the liquid was poured intro a petri dish with minimal agar and incubated for 48-72 hours at 37°C in the dark. Colonies (his+ revertants) were then counted.

Evaluation criteria:

Cytotoxicity: thinning of background lawn and reduction in colony numbers.
Genotoxicity: increase in revertant colony numbers

Statistics:

Not required.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

toxic to most strains at either 4 or 20 µg/plate and above

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not specified

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

toxic to most strains at either 4 or 20 µg/plate and above

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not specified

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

toxic to most strains at either 4 or 20 µg/plate and above

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not specified

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

toxic to most strains at either 4 or 20 µg/plate and above

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not specified

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

toxic to most strains at either 4 or 20 µg/plate and above

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

In Experiment 1 the test substance was toxic to most strains at either 4 or 20 µg/plate and above, both with and without metabolic activation. Therefore 100 µg/plate was selected was the highest dose level in Experiment 2.

The test compound did not cause a significant increase in the number of revertant colonies with any of the tester strains, neither in the absence or presence of S9 mix. No dose-dependent effect was obtained.

Sterility of the S9 mix and test compound were confirmed in sterility check plates.

Any other information on results incl. tables

No further information.

Applicant's summary and conclusion

Conclusions:

No evidence of mutagenicity was observed in either the presence or absence of metabolic activation.

Executive summary:

The mutagenic potential of Bardap 26 was evaluated in the bacterial reverse mutation assay (Ames Test). The test substance was applied to Salmonella typhimurium strains TA1535, TA1537, TA1538, TA98 and TA100, and Escherichia coli strain WP2uvrA, both in the presence and absence of metabolic activation (S9 mix). Test substance concentrations in the cytotoxicity test and first independent experiment were 0, 4, 20, 100, 500, 2500 and 10000 µg/plate. Cytotoxicity, evidenced by a thinning of the background lawn and a reduction in colony numbers, was observed in most strains with and without S9 at either 4 or 20 µg/plate and above. Therefore, test substance concentrations in the second independent experiment were reduced as follows: 0, 0.032, 0.16, 0.8, 4, 20 and 100 µg/plate. No significant increase in revertant colony numbers was observed, even at toxic dose levels, both with and without metabolic activation (S9 mix). It is concluded that the test substance is not mutagenic in the bacterial reverse mutation assay, and therefore does not require classification as a mutagen.