C16-18 (evennumbered, C18 unsaturated) alkyl bis(2-hydroxyethyl) amine oxide

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:

19 March 1990 - 3 April 1990

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study was performed under GLP and accoding to OECD/EC guidelines. In the current study the guidelines was not updated yet and this test does not contains: E. coli WP2 uvrA, or E. coli WP2 uvrA (pKM101), or S. typhimurium TA102. These strains are preferred to detect cross-linking mutagens.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Principles of method if other than guideline:

In the current study the guidelines was not updated yet and this test does not contains: E. coli WP2 uvrA, or E. coli WP2 uvrA (pKM101), or S. typhimurium TA102. These strains are preferred to detect cross-linking mutagens.

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

2,2'-(Z)-octadec-9-en-1-ylimino)bisethanol N-oxide

EC Number:

300-699-5

EC Name:

2,2'-(Z)-octadec-9-en-1-ylimino)bisethanol N-oxide

Cas Number:

93962-62-0

Molecular formula:

C22H45NO3

Details on test material:

Chemical name: Oleylbis(2-hydroxyethyl)amine oxide
Description: Light yellow liquid
Batch/Lot number: RCD/HJ-173
Purity: 58%
Instructions for test substance storage: In the original container at room temperature in the dark.
Stability under storage conditions: Stable
Expiry date: 09 March 1991

Method

Target gene:

Genes involved in histidine synthesis.

Species / strain

Species / strain / cell type:

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

Additional strain / cell type characteristics:

other: see remarks section

Metabolic activation:

with and without

Metabolic activation system:

S9-mix

Test concentrations with justification for top dose:

preliminary toxicity test (with and without S9-mix): 1.0, 3.3, 10.0, 33.3, 100, 333, 1000, 3330, 5000 µg/plate
experiment 1 without S9-mix: 0.33, 1.0, 3.3, 10.0, 33.3 µg/plate
experiment 1 with S9-mix: 1.0, 3.3, 10.0, 33.3, 100.0 µg/plate
experiment 2 without S9-mix: 0.33, 1.0, 3.3, 10.0, 33.3 µg/plate
experiment 2 with S9-mix: 1.0, 3.3, 10.0, 33.3, 100.0 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: not given, common solvent for this type of test.

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

dimtehylsulfoxide (DMSO)

Positive controls:

yes

Positive control substance:

other: see remarks section for details

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: none
- Exposure duration: 48 hours

NUMBER OF REPLICATIONS: each dose is tested in triplicate in each strain.

NUMBER OF CELLS EVALUATED: all colonies were counted.

DETERMINATION OF CYTOTOXICITY
- Method: other: Selection of an adequate range of doses was based on a preliminary toxicity test with strain TA100, both with and without S9-mix. Nine concentrations have been tested in duplicate for toxicity. The highest concentration of test article used in the subsequent mutagenesis assay was that which gave a reduced survival on the non-selective plates.

Evaluation criteria:

An Ames test was considered acceptable if it met the following criteria:
a) The negative control data (number of spontaneous revertants per plate) should reasonably fall within the laboratory background historical range
for each tester strain.
b) The positive control chemicals should produce responses in all tester strains which also reasonably fall within the laboratory historical range documented for each positive control substance. Furthermore, the mean plate count should be a t least two times the concurrent vehicle control group mean.
c) The selected dose range should include a clearly toxic concentration as demonstrated by the preliminary toxicity range-finding test with strain
TA100 or should extend to 5 mg/plate.

Statistics:

No formal hypothesis testing has been done.
A test substance was considered negative (not mutagenic) in the Ames test if:
a) The total number of revertants in any tester strain at any concentration was not greater than two times the solvent control value, with or
without metabolic activation.
b) The negative response should be reproducible in at least one independently repeated experiment.
A test substance was considered positive (mutagenic) in the Ames test if:
a) It induced at least a 2-fold, dose related increase in the number of revertants with respect to the number induced by the solvent control in any of the tester strains, either with or without metabolic activation. However, any mean plate count of less than 20 was considered to be not significant. If the test substance showed in the first test only a positive response at one or two concentrations, the assay was repeated with doses just below and exceeding those showing positive effects in the first test.
b) The positive response should be reproducible in at least one independently repeated experiment.
The preceding criteria were not absolute and other extenuating factors might enter into the final evaluation decision.

Results and discussion

Test results

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:

cytotoxicity

Remarks:

Without S9-mix 33.3µg/plate gives a moderate reduction and elimination at 100µg/plate. WithS9-mix with S9-mix 100µg/plate gives a moderate reduction and elimination at 333µg/plate.

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no data
- Effects of osmolality: no data
- Evaporation from medium: no data
- Water solubility: no data
- Precipitation: not observed
- Other confounding effects: no data


RANGE-FINDING/SCREENING STUDIES: The survival of the TA100culture i s determined by comparing the number of colonies on the plates containing the test substance with those on the solvent control plate. In the absence of S9-mix the survival of strain TA100 is moderately reduced at a test substance concentration of 33.3 µg/plate and eliminated at and above 100 µg/plate. In the presence o f S9-mix the survival of strain TA100 is moderately reduced at a test substance concentration of 100 µg/plate and eliminated at and above 333 µg/plate. Based on these data, the test substance was tested up to a concentration of 33.3 µg/plate i n the absence of S9-mix and up to 100 µg/plate in the presence of S9-mix.

COMPARISON WITH HISTORICAL CONTROL DATA: The negative and strain-specific positive control values fell within laboratory background historical ranges indicating that the test conditions were optimal and that the metabolic activation system functioned properly.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

The test substance did not induce a dose-related increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535; TA1537; TA98 and TA100) therefore from the results of this study it can be concluded that AROMOX O/12 is not mutagenic in the ames test.

Executive summary:

The test item was tested in the Ames Salmonella/microsome plate test up to 33.3 µg/plate in the absence o f S9 -mix and up to 100 µg/plate in the presence of S9 -mix. The test substance did not induce a dose-related increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535; TA1537; TA98 and TA100). These results were confirmed in an independently repeated experiment. The test item can, therefore, be considered as not mutagenic in this test system.