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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:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Not a GLP study
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
2010

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Deviations:
yes
Remarks:
No short-term exposure in absence of S9
Principles of method if other than guideline:
None
GLP compliance:
yes
Remarks:
The NTP conducts its studies in compliance with FDA Good Laboratory Practice Regulations
Type of assay:
in vitro mammalian chromosome aberration test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
- Physical state: Yellow liquid
- Stability under test conditions: Stable
- Storage condition of test material: Stored at or below - 20° C, protected from light, in 1-L Teflon® bottles.

Method

Target gene:
Not applicable
Species / strain
Species / strain / cell type:
Chinese hamster Ovary (CHO)
Details on mammalian cell type (if applicable):
- Type and identity of media: McCoy's 5A medium
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: No data
- Periodically checked for karyotype stability: No data
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S9
Test concentrations with justification for top dose:
0, 50, 100, 200, 300, 400, 500 without S9
0, 150, 160, 170, 180, 190, 200 with S9
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
no
Positive controls:
yes
Remarks:
Mitomycin C and Cyclophosphamide
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 10 hours without S9, 2 hours with S9
- Expression time (cells in growth medium): 2 hours without S9, 10 hours with S9
- Fixation time (start of exposure up to fixation or harvest of cells): 12 hours

SPINDLE INHIBITOR (cytogenetic assays): Colcemid
STAIN (for cytogenetic assays): Giemsa

NUMBER OF REPLICATIONS: 2

NUMBER OF CELLS EVALUATED: 200 per concentration

DETERMINATION OF CYTOTOXICITY
- Method: Presence of scorable metaphases
Rationale for test conditions:
Not reported
Evaluation criteria:
Chromosomal aberration data are presented as percentage of cells with aberrations. To arrive at a statistical call for a trial, analyses were conducted on both the dose response curve and individual dose points. For a single trial, a statistically significant (P#0.05) difference for one dose point and a significant trend (P#0.015) were considered weak evidence for a positive response; significant differences for two or more doses indicated the trial was positive. A positive trend test in the absence of a statistically significant increase at any one dose resulted in an equivocal call (Galloway et al., 1987). Ultimately, the trial calls were based on a consideration of the statistical analyses as well as the biological information available to the reviewers.
Statistics:
No data

Results and discussion

Test results
Key result
Species / strain:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
the high dose was limited by toxicity.
Vehicle controls validity:
not specified
Untreated negative controls validity:
not applicable
Positive controls validity:
not specified
Additional information on results:
Isoeugenol (in medium concentrations up to 200 µg/mL) did not induce chromosomal aberrations in cultured CHO cells, with or without S9 activation

Any other information on results incl. tables

See the attached document for information on tables of results

Applicant's summary and conclusion

Conclusions:
Isoeugenol (in medium concentrations up to 200 µg/mL) did not induce chromosomal aberrations in cultured CHO cells, with or without S9 activation.
Executive summary:

Testing was performed as reported by Gallowayet al.(1987). Isoeugenol was sent to the testing laboratory as a coded aliquot. It was tested in cultured Chinese hamster ovary (CHO) cells for induction of chromosomal aberrations (Abs), both in the presence and absence of Aroclor 1254-induced male Sprague Dawley rat liver S9 and cofactor mix. Cultures were handled under gold lights to prevent photolysis of bromodeoxyuridine-substituted DNA. Each test consisted of concurrent solvent and positive controls and of at least three doses of isoeugenol; the high dose was limited by toxicity. A single flask per dose was used. In the Abs test without S9, cells were incubated in McCoy’s 5A medium with isoeugenol for 10 hours; Colcemid was added, and incubation continued for 2 hours. The cells were then harvested by mitotic shake-off, fixed, and stained with Giemsa. For the Abs test with S9, cells were treated with isoeugenol and S9 for 2 hours, after which the treatment medium was removed and the cells were incubated for 10 hours in fresh medium, with Colcemid present for the final 2 hours. Cells were harvested in the same manner as for the treatment without S9. Cells were selected for scoring on the basis of good morphology and completeness of karyotype (21 ± 2 chromosomes). All slides were scored blind, and those from a single test were read by the same person. Two hundred first-division metaphase cells were scored at each dose level. Classes of aberrations included simple (breaks and terminal deletions), complex (rearrangements and translocations), and other (pulverized cells, despiralized chromosomes, and cells containing 10 or more aberrations).

Isoeugenol (in medium concentrations up to 200 µg/mL) did not induce chromosomal aberrations in cultured CHO cells, with or without S9 activation.