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EC number: 278-503-1 | CAS number: 76622-74-7
Genetic toxicity: in vitro
Administrative data
- Endpoint:
- in vitro cytogenicity / chromosome aberration study in mammalian cells
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 13 Jul 2009 - 19 Feb 2010
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 010
- Report date:
- 2010
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Landesamt für Umwelt, Wasserwirtschaft und Gewerbeaufsicht, Rheinland-Pfalz, Mainz, Germany
- Type of assay:
- in vitro mammalian chromosome aberration test
Test material
- Reference substance name:
- (Z)-N-methyl-N-(1-oxo-9-octadecenyl)glycine
- EC Number:
- 203-749-3
- EC Name:
- (Z)-N-methyl-N-(1-oxo-9-octadecenyl)glycine
- Cas Number:
- 110-25-8
- Molecular formula:
- C21H39NO3
- IUPAC Name:
- N-methyl-N-oleoylglycine
Constituent 1
Method
- Target gene:
- Not applicable
Species / strain
- Species / strain / cell type:
- lymphocytes: in whole blood treated with heparin
- Details on mammalian cell type (if applicable):
- RPMI 1640 medium supplemented with
- 15% (v/v) fetal calf serum (FCS)
- 1% (v/v) penicillin/streptomycin (10000 U /10000 µg/mL)
- 4.8 µg/mL Phytohaemagglutinin solution in H2O
During exposure to the test substance, RPMI medium was used without FCS supplementation.
- Metabolic activation:
- with and without
- Metabolic activation system:
- cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of rats treated with Aroclor 1254
- Test concentrations with justification for top dose:
- Pre-Experiment I:
4h treatment: 55.3, 110.6, 221.3, 442.5, 885, 1770 and 3540 µg/mL, with and without metabolic activation.
Pre-Experiment II:
4h treatment: 0.78, 1.56, 3.125, 6.25, 12.5, 25 and 50 µg/mL, without metabolic activation.
Experiment I:
4h treatment: 10, 15, 20, 25, 30, 35 and 40 µg/mL, with and without metabolic activation.
Experiment II:
4h treatment: 10, 15, 20, 25, 30, 35 and 40 µg/mL, with metabolic activation.
22h treatment: 0.5, 1, 5, 10, 15, 20, 25, 30, 40 an 50 µg/mL, without metabolic activation. - Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: DMSO
Controls
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- other: cyclophosphamide, 35 µg/mL in 0.9% NaCl, +S9; methanesulphonate, 600 and 350 µg/mL (experiment I and II, respectively) in nutrient medium, –S9
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in medium
DURATION
- Exposure duration: 4 and 22 h
- Fixation time (start of exposure up to fixation or harvest of cells): 22 h
SPINDLE INHIBITOR (cytogenetic assays): colchicine 0.1 pg/mL
STAIN (for cytogenetic assays): Giemsa 10% (v/v) in Soerensen buffer
NUMBER OF REPLICATIONS: 2
NUMBER OF CELLS EVALUATED: 100 per culture
DETERMINATION OF CYTOTOXICITY
- Method: mitotic index of 1000 cells
OTHER EXAMINATIONS:
- Determination of polyploidy: yes - Evaluation criteria:
- A test item is classified as non-mutagenic if: the number of induced structural chromosome aberrations in all evaluated dose groups is below 5.0 % aberrant cells, excluding gaps. No significant increase of the number of structural chromosome aberrations is observed.
A test item is classified as mutagenic if: the number of induced structural chromosome aberrations is above 5.0 % aberrant cells, excluding gaps and either a concentration-related or a significant increase in the number of cells with structural chromosome aberrations is observed. - Statistics:
- Statistical significance was calculated by means of the Fisher's exact test.
Results and discussion
Test results
- Species / strain:
- lymphocytes: in whole blood treated with heparin
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- from 30 µg/mL without and from 40 µg/mL with metabolic activation
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Additional information on results:
- TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: was observed from 221.3 µg/mL and above with and without metabolic activation.
RANGE-FINDING/SCREENING STUDIES: A preliminary cytotoxicity test was performed to determine the concentrations to be used in the mutagenicity assay. The applied concentrations ranged from 55.3 µg/mL to 3540 µg/mL with and without metabolic activation (exposure time 4 h, pre-experiment I). In pre-experiment II concentrations from 0.78 to 50 µg/mL were used.
ADDITIONAL INFORMATION ON CYTOTOXICITY: All concentraitons of the pre-experiment I showed cytotoxicity. - Remarks on result:
- other: strain/cell type: lymphocytes
- Remarks:
- Migrated from field 'Test system'.
Any other information on results incl. tables
Table 1. Results of cytogenicity experiments.
Test item |
Concentration |
Mitotic Index |
Aberrant cells in % |
||||
|
in µg/mL |
in % |
with gaps |
without gaps |
with exchanges |
||
|
Exposure period 4h, fixation time 22h, without S9 mix |
||||||
DMSO |
0.5% (v/v) |
100.0 |
5 |
1 |
0 |
||
EMS |
600 |
48.4 |
23 |
21.5* |
2.5 |
||
Test substance |
10 |
94.0 |
2.5 |
1.5 |
0 |
||
20 |
70.2 |
3 |
2 |
0.5 |
|||
30 |
50.4 |
0.5 |
0 |
0 |
|||
|
Exposure period 4h, fixation time 22h, with S9 mix |
||||||
DMSO |
0.5% (v/v) |
100 |
2 |
2 |
0 |
||
CPA |
35 |
77.2 |
21.5 |
16.5* |
0.5 |
||
Test substance |
15 |
82.6 |
2.5 |
2 |
0 |
||
30 |
39.1 |
2.5 |
1.5 |
0.5 |
|||
40 |
18.8 |
3 |
2 |
0 |
|||
|
Exposure period 22h, fixation time 22h, without S9 mix |
||||||
DMSO |
0.5% (v/v) |
100.0 |
0 |
0 |
0 |
||
EMS |
350 |
44.8 |
39.5 |
33.5* |
4.5 |
||
Test substance |
50 |
97.1 |
5.5 |
2 |
0 |
||
100 |
77.6 |
3.5 |
2 |
0 |
|||
125 |
36.8 |
2 |
1 |
0 |
|||
|
Exposure period 4h, fixation time 22h, with S9 mix |
||||||
DMSO |
0.5% (v/v) |
100.0 |
1.5 |
1.5 |
0 |
||
CPA |
35 |
70.1 |
33 |
22* |
1 |
||
Test substance |
25 |
94.8 |
3.5 |
2 |
0 |
||
35 |
69.7 |
3.5 |
3 |
0 |
|||
40 |
29.5 |
1 |
1 |
0 |
|||
*:Aberration frequency statistically significant higher than corresponding control values
CPA: Cyclophosphamide
EMS: Ethyl methanesulphonate
Applicant's summary and conclusion
- Conclusions:
- Intgerpretation of results: negative
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