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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
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given. Wrong positive control at 6 h exposure without S9 mix and highest test concentration of 2.2 mg/mL.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1998

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Deviations:
yes
Remarks:
wrong positive control at 6 h exposure without S9, highest test concentration 2.2 mg/mL.
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): Glycerol triacetate, Triacetin
- Analytical purity: 98.2 %

Method

Target gene:
Not applicable
Species / strain
Species / strain / cell type:
mammalian cell line, other: Chinese hamster lung (CHL/IU) cells
Metabolic activation:
with and without
Metabolic activation system:
cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of rats treated with phenobarbital and 5,6-benzoflavone
Test concentrations with justification for top dose:
Without metabolic activation
Continuous treatment: 0.55, 1.1 and 2.2 mg/mL
Short-term treatment: 0.55, 1.1 and 2.2 mg/mL

With metabolic activation
Short-term treatment: 0.55, 1.1 and 2.2 mg/mL
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: cyclophosphamide, 5 µg/mL, +S9; mitomycin C, 0.5 µg/mL, -S9
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: For continuous treatment, cells were treated for 24 or 48 h without S9.   
- Exposure duration: For short-term treatment, cells were treated for 6 h with and without S9 and cultivated with fresh media for 18 h.

NUMBER OF REPLICATIONS: 2 independent experiments were performed.

NUMBER OF CELLS EVALUATED: 200

DETERMINATION OF CYTOTOXICITY
- Method: Cell confluency was measured with MonocellaterTM

OTHER EXAMINATIONS:
- Determination of polyploidy: yes
Statistics:
Fisher's exact analysis.

Results and discussion

Test resultsopen allclose all
Species / strain:
mammalian cell line, other: CHL/IU
Metabolic activation:
without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
other: wrong positive control at 6 h exposure, valid at 24 and 48 h exposure
Species / strain:
mammalian cell line, other: CHL/IU
Metabolic activation:
with
Genotoxicity:
ambiguous
Remarks:
Effects at highest concentration were only observed at unphysiological pH value.
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at 2.2 mg/mL
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Structural chromosomal aberrations (including gaps) were induced on short-term treatment with an exogenous metabolic activation system (2.2 mg/mL). However, at this highest test substance concentration the medium colour was changed to yellow indicating a significant decrease of pH, accompanied by a high increase of cytotoxicity. These confounding factors do not allow an interpretation of the observed chromosomal damages as test-substance specific. It is known that acidification of the cell culture medium leads to chromosomal damages together with cytotoxicity in cell cultures. This is not expected to occur at similar test substance concentrations in vivo.
Polyploidy was not induced under any conditions.
Remarks on result:
other: strain/cell type:
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1. Chromosome analysis of Chinese hamster lung cells (CHL/IU) continuously treated with glycerol triacetate (TA) ** without S9 mix. 

 

Group

Concen-

tration

(mg/mL)

Exposure

(h)

No.

of

cells ana-lyzed

Cyto-

toxicity3

(%)

Mito-

tic index4

(%)

Aberrant cells in %

Poly-

ploid2

(%)

incl. gaps

excl. gaps

Non-treatment

 

 

200

-

-

0.0

0.0

0.25

Solvent1

0

24

200

100

-

0.5

0.5

0.5

Triacetin

0.55

24

200

100

-

1.0

1.0

0.0

Triacetin

1.1

24

200

92.5

-

1.5

1.5

0.25

Triacetin

2.2

24

200

90.5

8.6

0.0

0.0

0.0

MMC

0.00005

24

200

-

-

16.5*

15.5*

0.13

 

 

 

 

 

 

 

 

 

Solvent1

0

48

200

100

-

0.0

0.0

0.25

Triacetin

0.55

48

200

105

-

0.0

0.0

0.38

Triacetin

1.1

48

200

104

-

0.0

0.0

0.38

Triacetin

2.2

48

200

85

6.8

0.0

0.0

0.13

MMC

0.00005

48

200

-

-

20*

17*

0.25

1DMSO

2800 cells were analysed in each group

3Cell confluency, representing cytotoxicity, was measured with MonocellaterTM

4Number of metaphase per 500 cells was scored in each dish in order to select the highest dose enable to analyze chromosomes

*Significantly different from solvent control at p<0.01 by Fisher’s exact test

**Purity was 98.2% wt%. Diacetin (0.9%) was contained as impurity

MMC = Mitomycin C, positive control group

 

Table 2. Chromosome analysis of Chinese hamster lung cells (CHL/IU) treated with glycerol triacetate (TA)** with and without S9 mix. 

 

Group

S9 Mix

Concen-

tration

(mg/mL)

Expo-sure

(h)

No. of

cells ana-lyzed

Cyto-toxicity3

(%)

Mito-

tic index4

(%)

Aberrant cells in %

Poly-ploid2

(%)

incl. gaps

excl. gaps

Non-treatment

 

 

 

200

-

-

0.0

0.0

0.0

Solvent1

-

0

6-(18)

200

100

-

0.0

0.0

0.13

Triacetin

-

0.55

6-(18)

200

100.5

-

0.5

0.5

0.63

Triacetin

-

1.1

6-(18)

200

99.5

-

1.0

1.0

0.0

Triacetin

-

2.2

6-(18)

200

90.5

9.2

0.0

0.0

0.25

CPA

-

0.005

6-(18)

200

-

-

0.5

0.5

0.50

 

 

 

 

 

 

 

 

 

 

Solvent1

+

0

6-(18)

200

100

-

0.0

0.0

0.25

Triacetin

+

0.55

6-(18)

200

109

-

0.0

0.0

0.5

Triacetin

+

1.1

6-(18)

200

104.5

-

0.5

0.5

0.25

Triacetin

+

2.2***

6-(18)

200

25.5

1.6

43.5*

42.0*

0.26

CPA

+

0.005

6-(18)

200

-

-

45.5*

45*

0.13

1DMSO

2800 cells were analysed in each group

3Cell confluency, representing cytotoxicity, was measured with MonocellaterTM

4Number of metaphase per 500 cells was scored in each dish in order to select the highest dose enable to analyze chromosomes

*Significantly different from solvent control at p<0.01 by Fisher’s exact test

**Purity was 98.2 % wt%. Diacetin (0.9%) was contained as impurity

***Medium colour was changed to yellow, which showed lowering of pH, after treatment

CPA = Cyclophosphamide, positive control group

 

 

 

 

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative
Executive summary:

.