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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2004
Report Date:
2004

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Deviations:
no
Qualifier:
according to
Guideline:
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
Deviations:
no
GLP compliance:
yes
Type of assay:
in vitro mammalian chromosome aberration test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 2-amino-2-ethyl-1,3-propanediol
- Analytical purity: 99.4%

Method

Target gene:
not applicable
Species / strain
Species / strain / cell type:
other: Chinese hamster lung (CHL/IU) cells
Details on mammalian cell type (if applicable):
- Type and identity of media: Eagle´s MEM with 10 vol.% calf serum (deactivated)
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
cofactor supplemented post-mitochondrial fraction (S9-mix), prepared from the livers of rats treated with treated with phenobarbital and 5,6-benzoflavone
Test concentrations with justification for top dose:
without S9 mix(short-term treatment, continuous treatment 24 h and 48 h): 75, 150, 300, 600 and 1200 μg/mL
with S9 mix(short-term treatment): 75, 150, 300, 600 and 1200 μg/mL
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: physiol. saline
Controls
Untreated negative controls:
yes
Remarks:
medium
Negative solvent / vehicle controls:
yes
Remarks:
physiol. saline
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: without S9 mix: Mitomycin C (0.05 µg/mL), with S9 mix: Cyclophosphamide (15 µg/mL)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 6 h (short time treatment), 24 and 48 h (continuous treatment)
- Expression time (cells in growth medium): 18 h (for short time treatment)

SPINDLE INHIBITOR (cytogenetic assays): colcemide (final concentration: 0.2 µg/mL)
STAIN (for cytogenetic assays): Giemsa stain (2 vol.%)

NUMBER OF REPLICATIONS: 2 plates per test, 1 experiment

NUMBER OF CELLS EVALUATED: 200 metaphase images per dose (100 per plate)

DETERMINATION OF CYTOTOXICITY
- Method: The cell multiplication inhibiting action of the test substance was determined by measuring the cell density using a single-layer cultured cell densitometer and using the ratio of the cell density of the test group and of the negative group.
Evaluation criteria:
The judgment was made according to the criteria of Ishidate et al. (1987). When the frequencies of appearance of the cells with chromosomal aberrations (CA) were < 5%, the results were considered negative; when the frequencies of CA were 5% or > 5%, the results were considered false positive; when the frequencies of CA were 10% or greater, the results were considered positive. Finally, in the cases in which dose dependence or reproducibility was observed the appearance of aberrant cells, the results were considered positive.

Results and discussion

Test results
Key result
Species / strain:
other: Chinese hamster lung (CHL/IU) cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
RANGE-FINDING/SCREENING STUDIES:
Based on the results of performing cell multiplication inhibition tests, it was decided to use a test concentration range of 75-1200 µg/mL.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

No increase in chromosomal aberrations was observed in the test with either the short- term treatment (without S9 mix and with S9 mix) or continuous treatment.

Table 1: Results of experiments

Test item

Concentration

Cell growth ratio [mean in %]*

Aberrant cells in %

 

in µg/mL

in %

with gaps

without gaps

Exposure period 6-18 h, fixation time 24h, without S9 mix

saline

0.9%

100

2.5

1.0

MMC

0.05

79

25

24

Test substance

75

93

2.5

1.0

150

93

3.0

1.5

300

86

2.5

0.5

600

86

2.0

1.0

1200

79

2.0

1.5

Exposure period 6-18 h, fixation time 24h, with S9 mix

saline

0.9%

100

1.0

0

CP

15

93

79.0

79.0

Test substance

75

107

1.5

1.5

150

93

1.0

0.5

300

100

2.0

1.0

600

93

2.0

1.0

1200

85

2.5

1.5

Exposure period 24h, fixation time 24h, without S9 mix

saline

0.9%

100

4.0

2.0

MMC

0.05

89

43.5

42.0

Test substance

75

89

3.0

0.5

150

89

3.0

1.0

300

111

2.5

1.0

600

89

2.5

0.5

1200

67

4.9

3.7

Exposure period 48h, fixation time 48h, without S9 mix

saline

0.9%

100

3.0

0.5

MMC

0.05

94

68.0

66.5

Test substance

75

100

1.5

0

150

106

1.5

0.5

300

106

4.0

1.5

600

100

1.5

0.5

1200

50

4.8

3.6

MMC: Mitomycin C; CP: Cyclophosphamide (positive controls)

* The mean value showed as a growth ratio against the negative control value.

 

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative