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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro DNA damage and/or repair study
Remarks:
Type of genotoxicity: DNA damage and/or repair
Type of information:
experimental study
Adequacy of study:
key study
Study period:
No data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Test is performed similar to OECD Guideline 479; non GLP; Acceptable basic data.

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
2001

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 479 (Genetic Toxicology: In Vitro Sister Chromatid Exchange Assay in Mammalian Cells)
Deviations:
no
GLP compliance:
no
Type of assay:
sister chromatid exchange assay in mammalian cells

Test material

Constituent 1
Reference substance name:
peppermint oil
IUPAC Name:
peppermint oil
Details on test material:
- Name of test material (as cited in study report): essential oil extracted from peppermint (Menthaxpiperita L.)
- Composition of test material, percentage of components: menthol (59.17%), menthone (18.78%), L-limonene (5.16%), isomenthone (3.55%), ß-caryophyllene (2.93%), germacrene (1.73%), caryophyllene oxide (1.67%), bornyl acetate (1.08%), caraway aldehyde (0.61%), ß-pinene (0.61%), α-pinene (0.56%), myrcene (0.34%), eucalyptol (0.32%), isoeugenol (0.32%), methylcinemate (0.27%), sabinene (0.26%), ß-ocymene (0.25%).

Method

Target gene:
Not relevant
Species / strain
Species / strain / cell type:
lymphocytes: human whole peripheral blood culture
Details on mammalian cell type (if applicable):
- Type and identity of media: HEPES-buffered RPMI 1640 medium
Metabolic activation:
without
Metabolic activation system:
As whole blood cultures display many properties common with the liver microsomal cytochrome P450 system, no external metabolising enzymes were added.
Test concentrations with justification for top dose:
0.10, 0.15, 0.20, 0.25, and 0.30 µl/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: acetone 2.3 µl/ml
Controls
Untreated negative controls:
yes
Remarks:
untreated cells
Negative solvent / vehicle controls:
yes
Remarks:
acetone
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
Migrated to IUCLID6: 0.015 µl/ml
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 24 hours
- Fixation time (start of exposure up to fixation or harvest of cells): 24 hours

SPINDLE INHIBITOR (cytogenetic assays): Colchicine (0.5 ug/ml) for the last 3 hours of incubation
STAIN (for cytogenetic assays): Giemsa

NUMBER OF REPLICATIONS: Duplicate cultures for each treatment

NUMBER OF CELLS EVALUATED: No less than 25 second-mitotic division cells per culture were analysed

DETERMINATION OF CYTOTOXICITY
- Method: other: Cell replicative kinetics ((RI) Replicative Index). RI shows the average number of times cells have divided in culture.

OTHER: No less than 2 pilot experiments (using 2 to 3 concentrations) preceded 3 representative experiments.
Evaluation criteria:
No data
Statistics:
Statistical significance was confirmed by means of the Mann-Whitney U-test and the z-test (p < 0.05).

Results and discussion

Test results
Species / strain:
lymphocytes: human whole peripheral blood culture
Metabolic activation:
without
Genotoxicity:
negative
Remarks:
Relative weak SCE induction in a dose-independent manner
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
RANGE-FINDING/SCREENING STUDIES: 2 pilot experiments (using 2 to 3 concentrations) preceded 3 representative experiments. As in all cases good quantitative correspondence between pilot and main experiments was found, only pooled results of representative experiments were reported in this paper.

COMPARISON WITH HISTORICAL CONTROL DATA: No data

ADDITIONAL INFORMATION ON CYTOTOXICITY: Peppermint oil (Menthaxpiperita L.) clearly inhibited cell replicative kinetics. At concentrations of 0.20 ul/ml and above statistically significant inhibition of cell replicative kinetics was evident ((RI) Replicative Index).

Any other information on results incl. tables

Peppermint essential oil was a relative weak SCE inducer, it induced SCE in a dose-independent manner at cytotoxic concentrations. It seems that at high doses of Peppermint essential oil "saturation" of SCE-inducing capacity occurs.

 

Dose               SCE/cell ± S.E.M.

0.10 ul/ml       8.4 ± 0.4

0.15 ul/ml       10.2 ± 0.6

0.20 ul/ml       8.5 ± 0.3

0.25 ul/ml       10.1 ± 0.5

0.30 ul/ml       9.1 ± 0.4

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative without metabolic activation

Peppermint essential oil (Menthaxpiperita L.). was a relative weak SCE inducer, it induced SCE in a dose-independent manner at cytotoxic concentrations.
Executive summary:

A Sister Chromatid Exchange (SCE) test in vitro using human lymphocytes (whole peripheral blood cultures) was carried out on the essential oils extracted from peppermint (Menthaxpiperita L.). The test was performed similar to OECD Guideline 479. As whole peripheral blood cultures display many properties common with the liver microsomal cytochrome P450 system, no external metabolising enzymes were added. The cells were continuously exposed to peppermint oil at 0.10, 0.15, 0.20, 0.25, and 0.30 ul/ml for 24 hours. Cell replicative kinetics was determined by means of Replicative Index (RI).

Peppermint oil (Menthaxpiperita L.) clearly inhibited cell replicative kinetics. At concentrations of 0.20 ul/ml and above statistically significant inhibition of cell replicative kinetics was evident. Peppermint essential oil was a relative weak SCE inducer, it induced SCE in a dose-independent manner at cytotoxic concentrations. It seems that at high doses of Peppermint essential oil "saturation" of SCE-inducing capacity occurs.

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