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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:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1991
Report date:
1991
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report date:
1991
Reference Type:
review article or handbook
Title:
Unnamed
Year:
2008

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: OECD 473 (1983)
Deviations:
yes
Remarks:
the stability of the test substance has not been reported; the stability and achieved concentration of the test substance in the vehicle used were not determined by analysis
Principles of method if other than guideline:
Method: according to OECD Guide-line 473 (1983) and UKEMS Recommended Procedure for Basic Mutagenicity Tests (Kirkland, 1990)
GLP compliance:
yes
Type of assay:
other: cultured human lymphocytes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Supplier: ICI Chemicals & Polymers Limited, Runcorn, UK
- Expiration date of the lot/batch: no data
- Stability under test conditions: stable
- Storage condition of test material: in the fridge, in the dark

Method

Species / strain
Species / strain / cell type:
other: human lymphocytes
Metabolic activation:
with and without
Metabolic activation system:
AROCLOR 1254 induced rat liver microsomal fraction S9 mix
Test concentrations with justification for top dose:
Range-finding cytotoxicity test:
8, 40, 200, 1000 and 5000 µg/mL
Main cytogenetic tests:
-S9-mix: 25, 100, 200, 400, 600, and 800 µg/mL
+S9-mix: 100, 500, 1000, 2000, 3500 and 5000 µg/mL
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO, Dimethylsulphoxide
- Justification for choice of solvent/vehicle: The solvent was chosen according to its solubility properties.
Controlsopen allclose all
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
Dosing solutions prepared in sterile double deionised water. Migrated to IUCLID6: Final concentration: 1.0 µg/mL
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
Dosing solutions prepared in sterile double deionised water. Migrated to IUCLID6: Final concentration: 50 µg/mL
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Preincubation period:
- Exposure duration: 72 and 96 hours
- Expression time (cells in growth medium): 3 hours
- Fixation time (start of exposure up to fixation or harvest of cells): at 72 or 96 hours

SPINDLE INHIBITOR (cytogenetic assays): colcemid (0.4 µg/mL)
STAIN (for cytogenetic assays): Giemsa and mounted in DPX

NUMBER OF REPLICATIONS:2

NUMBER OF CELLS EVALUATED: 100 cells per culture were evaluated (examination of cells with abberations); 1000 lymphocytes per culture (examination of mitose index)

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index
Statistics:
Evaluation was performed only for cells carrying aberrations exclusive gaps. The cells for each of the treatment groups were compared to the solvent control values using the Fisher's Exact Test (one sided).

Results and discussion

Test results
Species / strain:
lymphocytes: human lymphocytes
Metabolic activation:
without
Genotoxicity:
positive
Remarks:
at 600 µg/mL
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at 1000 and 5000 µg/mL in the absence of S9 or at 5000 µg/mL in the presence of S9-mix.
Vehicle controls validity:
valid
Positive controls validity:
valid

Any other information on results incl. tables

Results:
Increases in the percentage of aberrant cells were observed at 100, 400, and 600 mg/l in male donor cultures at 72 h sampling time and in female donor cultures at 600 mg/l harvested after 72 h; no significant increases were observed
at 96 h sampling time.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
positive in vitro in the absence of S9-mix

In the Chromosomal aberration test with human lymphocytes according to OECD 473 in the absence of mammalian activation S9-mix structural chromosomal aberrations were observed. Therefore, Ethyleneglycol dimethacrylate is considered to be clastogenic (in vitro in the absence of S9-mix) at 600 µg/mL under the experimental conditions reported.
Executive summary:

In a cytogenetic assay (Chromosome aberration test with human lymhocytes from two donors (1 f, 1 m) according to OECD 473) human lymphocytes were exposed to Ethylene glycol dimethacrylate (purity: 98 % w/w, solvent: DMSO) at concentrations in the range of 25 to 5000 µg/mL

in the presence and absence of mammalian metabolic activation S9 -mix (Aroclor 1254 induced rat liver homogenate). 

 

The assay was performed in two independent experiments using Ethylene glycol dimethacrylate (EGDMA) concentrations of 25, 100, 200, 400, 600 and 800 µg/mL in the presence of S9 -mix and concentrations of 100, 500, 1000, 2000 , 3500 and 5000 µg/mL in the absence of S9 -mix. Dose related reductions in mitotic activity were observed in cultures from both donors in the presence and absence of S9 -mix at the 72 hour sampling time.

Cultures treated with higher concentrations of EGDMA (600 µg/mL -S9 -mix and 1000 µg/mL +S9 -mix) in the main cytogenetic tests were considered not to be suitable for chromosomal aberration analysis due to lack of metaphases as a result of toxicity or due to cytotoxic effects on the chromosome structure.

Cultures from both donors exposed with EGDMA at concentrations of 100, 400 and 600 µg/mL in the absence of S9 -mix and 100, 500 and 1000 µg/mL in the presence of S9 -mix were choosen for chromosomal aberration analysis at 72 hours sampling time. Additionally, cultures from the female donor treated at 400 µg/mL in the absence of S9 -mix and 1000 µg/mL in the presence of S9 -mix were selected for analysis at 96 hour sampling time.

No statistically or biologically significant increases in percentage of aberrant cells, compared to the solvent control values, were seen at any of the EGDMA concentrations tested in either donor, in the presence of S9 -mix at the 72 hour sampling time or in the presence or absence of S9 -mix at 96 hour sampling time. Statistically significant increases of chromosomal aberrations were seen in cultures from both donors treated with EGDMA at 600 µg/mL in the absence of S9 -mix and examined at 72 hours sampling time.

No significant increases were observed at the 96-hours.

Positive controls (mitomycin C and cyclophosphamide) induced the appropriate response confirming the sensitivity of the test system. 

Therefore EGDMA showed under the experimental conditions reported reproducibly induced chromosomal damage in human peripheral blood lymphocytes in vitro in the absence of auxiliary mammalian metabolic activation S9 -mix.

This study is classified as acceptable. This study satisfies the requirement for Test Guideline OECD 473 for in vitro cytogenetic mutagenicity data. 

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