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Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information
Alpha-Terpineol gave negative results in the Ames test with Salmonella typhimurium strains TA98, TA100, TA1535, TA1537, and TA 1538 and in the in vitro mouse lymphoma assay, both with and without metabolic activation. Negative results were observed for its read-across candidate Terpineol multi (a multi-constituent substance with alpha-Terpineol as its main constituent and gamma-Terpineol as the minor constituent), in the in vitro chromosome aberration assay both with and without metabolic activation. Based on these results, alpha-Terpineol is considered not to be genotoxic.
Link to relevant study records
Reference
Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
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: Non GLP study but conducted similarly to OECD guideline 471.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
yes
Remarks:
no details about test substance and solvent or vehicle control
GLP compliance:
no
Type of assay:
bacterial reverse mutation assay
Target gene:
histidine
Species / strain / cell type:
S. typhimurium, other: TA98, TA100, TA1535, TA1537 and TA1538
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
Liver S9 prepared from male Sprague-Dawley rats and Syrian golden hamsters injected with Aroclor 1254 at 500 mg/kg body weight
Test concentrations with justification for top dose:
Ranged between 10 µg/plate and 1000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: no data
- Justification for choice of solvent/vehicle: no data
Untreated negative controls:
no
Negative solvent / vehicle controls:
not specified
True negative controls:
no
Positive controls:
yes
Positive control substance:
congo red
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium; in agar (plate incorporation) and preincubation


DURATION
- Preincubation period: 30 min
- Exposure duration: 48 h
Evaluation criteria:
Test article had to induce doubling the mean number of revertants / plate
Statistics:
No details given in study report
Species / strain:
S. typhimurium, other: TA98, TA100, TA1535, TA1537 and TA1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
not specified
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Conclusions:
Interpretation of results (migrated information):
negative

Alpha-terpineol was not mutagenic in the Ames test in both plate incorporation and preincubation methods with and without metabolic activation.
Executive summary:

In a reverse gene mutation assay in bacteria conducted similarly to OECD guideline 471, TA98, TA100, TA1535, TA1537 and TA1538 strains of S. typhimurium  were exposed to Alpha-terpineol at concentrations between 1µg and 1000 µg/plate in the presence and absence of mammalian metabolic activation system liver S9 homogenate, from male Sprague-Dawley rats and Syriyan golden hamsters injected with Aroclor 1254 at 500 mg/kg body weight.

Alpha-terpineol was tested for mutagenicity at different dose concentrations with both direct plate incorporation and preincubation methodology. Alpha-terpineol caused no dose-related response in the number of histidine auxotroph revertants. The positive controls induced the appropriate responses in the corresponding strains.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vitro:

For alpha-Terpineol an Ames test and an in vitro gene mutation test with mammalian cells are available but not a cytogenicity assay. To cover the latter endpoint an in vitro chromosomal aberration test performed with Terpineol multi (a multi-constituent substance with alpha-Terpineol as its main constituent and gamma-Terpineol as the minor constituent), will be used for read-across. In the toxico-kinetic section) the constituents of alpha-Terpineol and Terpineol multi are presented. These terpineols are expected to have a similar genotoxicity profile because of their similarity in structure. It can also be seen that all constituents of Terpineol multi have a similar backbone, which is the cyclohexyl ring. The attached methyl groups are para-positioned. There are two functional groups. The first one is the tertiary alcohol, which is not reactive because no additional reactive groups are adjacent to this alcohol. The second functional group is the double bond, at the para–position but can be inside or outside the cyclohexyl ring. These differences are thought to be of minor importance for the genotoxicity potential

Alpha-Terpineol: Ames test

In a reverse gene mutation assay in bacteria conducted similarly to OECD guideline 471,TA98, TA100, TA1535, TA1537 and TA1538 strains of S. typhimurium were exposed to alpha-Terpineol at concentrations between 1µg and 1000 µg/plate in the presence and absence of mammalian metabolic activation system liver S9 homogenate, from male Sprague-Dawley rats and Syriyan golden hamsters injected with Aroclor 1254 at 500 mg/kg body weight (Seifried 2006). Alpha-Terpineol was tested for mutagenicity at different concentrations with both direct plate incorporation and preincubation methodology. Alpha-Terpineol caused no dose-related response in the number of histidine auxotroph revertants. The positive controls induced the appropriate responses in the corresponding strains.

Alpha-Terpineol MLA study

In a mammalian cell gene mutation assay conducted similarly to OECD guideline 476, mouse lymphoma L5178Y cells cultured in vitro were exposed to alpha-Terpineol at concentrations between 0.14 µg/mL and 0.65 µg/mL in the presence and absence of metabolic activation with liver S9 prepared from Aroclor 1254-induced male Sprague-Dawley rats (Seifried 2006). Alpha-Terpineol was tested for cytotoxic concentration up to an upper limit of 10000 µg/plate. In both non-activated and S9-activated conditions, response was negative at a dose 0.14-0.65 µg/mL. The positive controls ethylmethylsulfonate (without metabolic activation) and 3-methylcholanthrene (with metabolic activation) induced the appropriate response.

Terpineol multi: Chromosome aberration study

In an in vitro chromosome aberration test performed according to OECD guideline 473 and in compliance with GLP, human primary lymphocyte cultures were exposed to Terpineol multi in DMSO at concentration range of 5.598-1543 μg/mL, for 3 + 17 h (treatment + recovery) with metabolic activation (2% S-9 fraction of Aroclor 1254-induced male Sprague-Dawley rats), and for 3 + 17 h or 20 + 0 h (treatment + recovery) without metabolic activation for a preliminary cytotoxicity test (Lloyd 2010). In the main test, two experiments were performed at concentrations up to 600 µg/mL without S-9 and up to 800 µg/mL with S-9 and the following concentrations were selected for analysis: Experiment 1: Without S-9 (treatment: 3 h): 0, 350, 425 and 450 μg/mL; with S-9 (treatment: 3 h): 0, 300, 550 and 625 μg/mL. Experiment 2: Without S-9 (treatment: 20 h): 0, 75, 200 and 225 μg/mL; with S-9 (treatment: 3 h): 0, 400, 550, 625 and 650 μg/mL. Proportion of cells with structural aberrations in negative control cultures fell within historical vehicle control ranges. Positive controls (4-nitroquinoline-N-oxide at 2.5 and 5 µg/mL without S-9 and cyclophosphamide at 10, 20 and 30 µg/mL with S-9) induced the appropriate response. Treatment of cells with Terpineol multi in the presence or absence of S-9 in both experiments resulted in frequencies of cells with structural or numerical aberrations that were generally similar to those observed in concurrent vehicle controls for all concentrations analysed. Numbers of aberrant cells (excluding gaps) in treated cultures fell within the normal range with the exception of one culture at the highest concentration analysed with S-9 in experiment 1 (625.0 µg/mL). However, the aberration frequency (excluding gaps) in the replicate culture at 625.0 µg/mL in experiment 1 and in all other cultures analysed in experiments 1 and 2 fell within the normal range. Under the test conditions, Terpineol multi is not considered as clastogenic in human lymphocytes.


Justification for selection of genetic toxicity endpoint
For finalising a conclusion on the endpoint genotoxicity all three standard in vitro tests are needed. The Ames test is considered to be the most predictive for genotoxicity and therefore, this has been selected here.

Justification for classification or non-classification

Based on the negative results of the Ames test and the mouse lymphoma assay with alpha-Terpineol and the negative result of the in vitro chromosome aberration assay with Terpineol multi (a multi-constituent substance with alpha-Terpineol as its main constituent and gamma-Terpineol as the minor constituent) alpha-Terpineol does not need to be classified for genotoxicity in vitro according to EU Directive 67/548 (DSD) and EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.