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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2011-11-03 to 2012-01-18
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
N,N-dimethyldecylamine N-oxide
EC Number:
220-020-5
EC Name:
N,N-dimethyldecylamine N-oxide
Cas Number:
2605-79-0
Molecular formula:
C12H27NO
IUPAC Name:
decyl(dimethyl)amine oxide
Test material form:
other: aqueous solution
Details on test material:
- Name of test material (as cited in study report): N,N-dimethyldecylamine-N-oxide (solution)
- Substance type: colourless to yellowish liquid
- Analytical purity: 40.5% active
- Impurities (identity and concentrations): peroxide: 0.01%, free amine: < 0.5%
- Purity test date: 2011-11-14
- Lot/batch No.: 1108042301
- Expiration date of the lot/batch:2 years in original closed packaging
- Storage condition of test material: 10 - 25°C

Method

Target gene:
his- → his+ reversion
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Metabolic activation system:
Arochlor 1254 induced rat liver S9
Test concentrations with justification for top dose:
3.16, 10.0, 31.6, 100, 316 and 1000 µg per plate.
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: completely soluble in water and represents a standard solvent in the testing facility.
Controls
Untreated negative controls:
yes
Remarks:
DMSO (vehicle)
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
no
Positive controls:
yes
Remarks:
-S9: TA 98, TA 102, TA 1537 - 2-amino-anthracene in DMSO (2 µg/plate); TA 100, TA 1535 - cyclophosphamide in aqua ad iniectabilia (1500 µg/plate)
Positive control substance:
other: +S9: TA 1535 and TA 100 - sodium azide in aqua ad iniectabilia (10 µg/plate); TA 98 - 2-nitro-fluorene in DMSO (10 µg/plate); TA 1537 - 9-amino-acridine in ethanol, abs. (100 µg/plate); TA 102 - methyl methane sulfonate (MMS) in DMSO (1300 µg/pl
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 2nd test only: 20 minutes
- Exposure duration: 48 - 72 hours
- Expression time (cells in growth medium): 10 hours

NUMBER OF REPLICATIONS: 3

NUMBER OF CELLS EVALUATED: 10E08

DETERMINATION OF CYTOTOXICITY
- Method: 50% reduction in colony number and scarce background lawn growth
Evaluation criteria:
The AMES test data are evaluated using the statistical method described below. A response is considered positive if
- the number of revertants is significantly increased (p =< 0.05, U-test according to MANN and WHITNEY) compared with the solvent control to at least 2-fold of the solvent control for TA 98, TA 100 and TA 102 and 3-fold of the solvent control for TA 1535 and TA 1537 in both independent experiments;
- in addition, a significant (p =< 0.05) concentration (log value)-related effect (Spearman’s rank correlation coefficient) is observed;
- positive results have to be reproducible and the histidine independence of the revertants has to be confirmed by streaking random samples on histidine-free agar plates.
The range of spontaneous reversion frequencies in our laboratory is generally:

TA 98: 20 - 60
TA 100: 100 - 200
TA 102: 240 - 320
TA 1535: 10 - 35
TA 1537: 3 - 20

The numbers may be slightly different on plates with S9 and may vary slightly from experiment to experiment.
Cytotoxicity is defined as a reduction in the number of colonies by more than 50% compared with the solvent control and/or a scarce background lawn.
Statistics:
See above.

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
1000 µg/plate all test strains, with/without activation
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

See attached tables

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

No mutagenic effect (no significant increase in revertant colony numbers as compared with control counts) was observed for N,N-dimethyldecylamine-N-oxide (solution), tested up to a cytotoxic dose level of 1000 µg N, N-dimethyl-decylamine-N-oxide/plate, in any of the 5 test strains in two independent experiments without and with metabolic activation, respectively (plate incorporation and preincubation test).
Executive summary:

In an in vitro reverse gene mutation (Ames) study performed in accordance with OECD Guideline 471 and EC Method B.13/14,

N,N-dimethyldecylamine-N-oxide (solution) was examined in the 5 Salmonella typhimurium strains TA 98, TA 100, TA 102, TA 1535 and TA 1537 in two independent experiments, each carried out without and with metabolic activation (a microsomal preparation derived from Aroclor 1254-induced rat liver). The first experiment was carried out as a plate incorporation test and the second as a preincubation test.

N,N-dimethyldecylamine-N-oxide (solution) was completely dissolved in dimethyl-sulfoxide (DMSO). A correction factor of 2.47 was used to correct for the content of active matter in the supplied solution (40.5%).

Ten concentrations ranging from 0.316 to 5000 µg N,N-dimethyldecylamine-N-oxide/plate were tested in two preliminary cytotoxicity tests (plate incorporation test without and with metabolic activation) in test strain TA 100. Pronounced cytotoxicity (scarce background lawn and reduction of the number of revertants by more than 50%) was noted in the plate incorporation test without and with metabolic activation at concentrations of 1000 µg/plate and higher.

Hence, 1000 µg N,N-dimethyldecylamine-N-oxide/plate was chosen as the maximum dose level for the main study in the plate incorporation test and in the preincubation test. Six concentrations ranging from 3.16 to 1000 µg N,N-dimethyldecylamine-N-oxide per plate were employed in two independent experiments each carried out without and with metabolic activation.

In two independent plate incorporation and preincubation tests, each carried out without and with metabolic activation, cytotoxicity (scarce background lawn and reduction of the number of revertants) was noted at the maximum dose level of 1000 µg/plate in all test strains

No mutagenic effect (no increase in revertant colony numbers as compared with control counts) was observed for the test material, tested up to the cytotoxic dose level of 1000 µg in any of the 5 test strains.

Under the present test conditions the test material tested up to a cytotoxic dose level of 1000 µg N,N-dimethyldecylamine-N-oxide/plate, caused no mutagenic effect in the Salmonella typhimurium strains TA 98, TA 100, TA 102, TA 1535 and TA 1537 neither in the plate incorporation test nor in the preincubation test each carried out without and with metabolic activation.

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