Tetramethylene dimethacrylate

Genetic toxicity: in vitro

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

Study period:

1999-11-10 to 1999-12-16

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

Report date:

2000

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

adopted 21 July 1997

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EEC Council Directive 92/69, part B

Deviations:

no

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

Tetramethylene dimethacrylate

EC Number:

218-218-1

EC Name:

Tetramethylene dimethacrylate

Cas Number:

2082-81-7

Molecular formula:

C12H18O4

IUPAC Name:

butane-1,4-diyl bis(2-methylacrylate)

Test material form:

liquid

Specific details on test material used for the study:

- Name of test material (as cited in study report): 1,4-Butanediol dimethacrylate

Method

Target gene:

not applicable (chromosome aberration test)

Species / strain

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

- Type and identity of media: Ham's F10 + 15% FCS
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
- Periodically "cleansed" against high spontaneous background: no
- other: generation time, plating efficiency checked

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

1250, 625, 313, 156, 78.1, 39.1, 19.5 and 9.77 mg/L

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: not given

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

with metabolic activation

Controls 2

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

without metabolic activation

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 3 h; second experiment: 20 h continuous exposure without S9 mix
- Expression time (cells in growth medium): 17 h (recovery period)
- Fixation time (start of exposure up to fixation or harvest of cells): 20 h

SELECTION AGENT (mutation assays):
SPINDLE INHIBITOR (cytogenetic assays): 0.2 µg/mL colcemid for the last 3 h of the treatment period
STAIN (for cytogenetic assays): 3% Giemsa

NUMBER OF REPLICATIONS: 2

NUMBER OF CELLS EVALUATED: 100 from each culture, two cultures were prepared at each test point

DETERMINATION OF CYTOTOXICITY
- Method: cell count

OTHER EXAMINATIONS:
- Determination of polyploidy: yes, recorded, but not included in the count of eligible metaphases
- Determination of endoreplication: yes, recorded, but not included in the count of eligible metaphases

Evaluation criteria:

A substance is considered clastogenic if: - any dose level shows a statistically significant increase in aberration-bearing cells - the increase is over historical controls - the increase is present in both replicates

Statistics:

Fisher's exact test with solvent control as reference point

Results and discussion

Test results

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no effects
- Effects of osmolality: no effects
- Precipitation: opacity occurred above 313 mg/L

COMPARISON WITH HISTORICAL CONTROL DATA:
- the number of cells with chromosome aberrations were in the range of the historical control data

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- in the first experiment in the absence of S9 mix cytotoxicity was observed at 625 mg/L and higher (number of viable cells reduced to 3 and 13% compared to solvent control); at 313 mg/L the number of viable cells was 61% compared to solvent control
- in the second experiment marked toxicity was observed at 313 mg/L and higher (cell counts < 22% compared to solvent control); at 156 mg/L the number of viable cells was reduced to 54% of the solvent control
- in the presence of S9 mix, no dose-related toxicity was observed

Remarks on result:

other: all strains/cell types tested

Any other information on results incl. tables

The test substance did not induce an increase in chromosomal aberrations at any test point selected for scoring in the absence or presence of S9 mix.

A slight, but not dose-related increase in endoreduplication, was observed in the presence of S9 mix.

Increases in gaps over the negative control value were observed in the 3 h treatment group (313 mg/L) without metabolic activation. This was seen in only one replicate and therefore not considered biologically relevant.

Significant increase in aberration-bearing cells was observed in the positive controls indicating the correct functioning of the test system.

3 h treatment, without S9 mix	mg/L	mean rel. Cell count %	cells scored	cells with aberrations (+gaps)	cells with aberrations (-gaps)	%CA
untreated		107	200	11	4	2.0
solvent		100	200	5	3	1.5
test substance	1250	13				–
	625	3				–
	313 (#)	61	200	15	5	2.5
	156 (#)	93	200	8	4	2.0
	78.1 (#)	79	200	12	6	3.0
	39.1	87				–
	19.5	101				–
	9.77	94				–
Mitomycin C	0.3	75	100	72	68	68.0 ***
3 h treatment, with S9 mix	mg/L	mean rel. Cell count %				%CA
untreated		70	200	9	3	1.5
solvent		100	200	11	4	2.0
test substance	1250 (#)	89	200	16	6	3.0
	625 (#)	69	200	14	8	4.0
	313 (#)	103	200	19	7	3.5
	156	86				–
	78.1	68				–
	39.1	86				–
	19.5	107				–
	9.77	69				–
Cyclophosphamide	15	88	100	62	56	56.0 ***
20 h treatment, without S9 mix	mg/L	mean rel. Cell count %				%CA
untreated		143	200	1	0	0.0
solvent		100	200	3	0	0.0
test substance	1250	0				–
	625	3				–
	313	22				–
	156 (#)	54	200	6	2	1.0
	78.1 (#)	112	200	3	1	0.5
	39.1 (#)	116	200	7	1	0.5
	19.5	120				–
	9.77	94				–
Mitomycin C	0.3	55	150	62	62	41.3 ***

%CA = percentage of cells bearing aberrations (excl. gaps)

(#) used for scoring

* statistically significant at p< 0.05

** statistically significant at p< 0.01

*** statistically significant at p< 0.001

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

In this chromsome aberration test 1,4-BDDMA did not induce chromosomal aberrations in CHO cells after in vitro treatment in the presence and absence of S9 metabolic activation.

Executive summary:

In a mammalian cell chromosome aberration assay according to OECD guideline 473, adopted 21 July 1997, CHO cell cultures were exposed to 1,4-BDDMA (93.59% a.i.) in DMSO at concentrations of 0 (control), 1250, 625, 313, 156, 78.1, 39.1, 19.5 and 9.77 mg/L with and without metabolic activation (S9 mix). 1,4-Butanediol dimethacrylate was tested up to cytotoxic concentrations.

In the first experiment the cells were exposed to the test substance for 3 h with and without metabolic activation and in a second experiment for 20 h without metabolic activation.

The test substance did not induce an increase in chromosomal aberrations at any test point selected for scoring in the absence or presence of S9 mix.

A slight, but not dose-related increase in endoreduplication, was observed in the presence of S9 mix.

Increases in gaps over the negative control value were observed in the 3 h treatment group (313 mg/L) without metabolic activation. This was seen in only one replicate and therefore not considered biologically relevant.

Positive controls induced the appropriate response.

There was no evidence of chromosome aberrations induced over background.

NOTE: Any of data in this dataset are disseminated by the European Union on a right-to-know basis and this is not a publication in the same sense as a book or an article in a journal. The right of ownership in any part of this information is reserved by the data owner(s). The use of this information for any other, e.g. commercial purpose is strictly reserved to the data owners and those persons or legal entities having paid the respective access fee for the intended purpose.