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EC number: 202-796-7 | CAS number: 99-87-6
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Genetic toxicity: in vitro
Administrative data
- Endpoint:
- in vitro gene mutation study in bacteria
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2020-06-22 to 2020-08-27
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 020
- Report date:
- 2020
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
- Version / remarks:
- 2008
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Version / remarks:
- 2020
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
Test material
- Reference substance name:
- p-cymene
- EC Number:
- 202-796-7
- EC Name:
- p-cymene
- Cas Number:
- 99-87-6
- Molecular formula:
- C10H14
- IUPAC Name:
- 1-isopropyl-4-methylbenzene
Constituent 1
Method
- Target gene:
- The Salmonella typhimurium histidine (his) and the Escherichia coli tryptophan (trp) reversion system measures his- → his+ and trp- → trp+ reversions, respectively. The strains are constructed to differentiate between base pair (E.coli WP2, TA 1535, TA 100) and frameshift (TA 1537, TA 98) mutations.
Species / strainopen allclose all
- Species / strain / cell type:
- E. coli WP2 uvr A
- Additional strain / cell type characteristics:
- not applicable
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
- Additional strain / cell type characteristics:
- not applicable
- Metabolic activation:
- with and without
- Metabolic activation system:
- Type and composition of metabolic activation system:
- Source of S9: Phenobarbital/β-naphthoflavone induced rat liver
- Method of preparation of S9 mix: The S9 was prepared and stored according to the currently valid version of the SOP for rat liver S9 preparation
- Concentration or volume of S9 mix and S9 in the final culture medium: 50 µl S9 and 500 µL S9 mix in the final culture medium (2700 µl total)
- Quality controls of S9 (e.g., enzymatic activity, sterility, metabolic capability): Each batch of S9 was routinely tested for its capability to activate the known mutagens benzo[a]pyrene and 2-aminoanthracene in the Ames test - Test concentrations with justification for top dose:
- Experiment I (with and without S9 mix): 3, 10; 33; 100; 333; 1000; 2500 and 5000 μg/plate;
Experiment II (with and without S9 mix):
- Strain TA98: 3, 10; 33; 100; 333; 1000; 2500 and 5000 μg/plate
- All remaining strains: 10; 33; 100; 333; 1000; 2500; and 5000 μg/plate
Justification for top dose: tested up to recommended maximum test concentration according to OECD 471 - Vehicle / solvent:
- - Vehicle/solvent used: DMSO (test item, positive controls),deionised water (positive controls)
- Justification for choice of solvent/vehicle: The solvent was chosen because of its solubility properties and its relative nontoxicity to the bacteria (Maron et al.; 1981).
Controlsopen allclose all
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- other: 2-aminoanthracene
- Remarks:
- - with S9
- 2.5 µg/plate (TA 1535, TA 1537, TA 98, TA 100) and 10 µg/plate (WP2 uvrA)
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Remarks:
- deionised water
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- methylmethanesulfonate
- Remarks:
- - without S9
- 2 µg/plate (WP2 uvrA)
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- other: 4-nitro-o-phenylene-diamine, 4-NOPD
- Remarks:
- - without S9
- 10 µg/plate (TA98), 50 µg/plate (TA1537)
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Remarks:
- deionised water
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- Remarks:
- - without S9
- 10 µg/plate (TA1535 and TA100)
- Details on test system and experimental conditions:
- NUMBER OF REPLICATIONS:
- Number of cultures per concentration: Triplicate
- Number of independent experiments: 2
METHOD OF TREATMENT/ EXPOSURE:
- Experiment I: Plate incoporation method
- Experiment II: Pre-incubation method
TREATMENT AND HARVEST SCHEDULE:
- Preincubation period: 60 minutes
- Duration of treatment: For at least 48 hours
METHODS FOR MEASUREMENT OF CYTOTOXICITY
- Method: Reduction in the number of spontaneous revertants (below a factor of 0.5) or a clearing of the bacterial background lawn
METHODS FOR MEASUREMENTS OF GENOTOXICIY
- Counting of revertant colonies using a Petri Viewer Sorcerer Colony Counter 3.0 (Instem, Suffolk IP33 3TA, UK) with the software program Ames Study Manager (v1.24) and Ames Archive Manager (v1.01). - Rationale for test conditions:
- In experiment I, the concentration range of the test item was 3 – 5000 μg/plate. Since toxic effects were observed in experiment 1, seven concentrations were tested in experiment II, except of strain TA 98, in which eight concentrations were tested. 5000 μg/plate were chosen as maximal concentration (recommended maximum test concentration according to OECD 471). The concentration range included two logarithmic decades.
Precipitation:
- from 1000 to 5000 μg/plate in the experiment I (in the overlay agar)
- from 2500 to 5000 μg/plate in experiment II
- at 5000 μg/plate in the overlay agar in both experiments - Evaluation criteria:
- 1) A test item is considered as a mutagen if a biologically relevant increase in the number of revertants of twofold or above (strains TA 98, TA 100, and WP2 uvrA) or threefold or above (strains TA 1535 and TA 1537) the spontaneous mutation rate of the corresponding solvent control is observed.
2) A dose dependent increase is considered biologically relevant if the threshold is reached or exceeded at more than one concentration.
An increase of revertant colonies equal or above the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.
3) A dose dependent increase in the number of revertant colonies below the threshold is regarded as an indication of a mutagenic potential if reproduced in an independent second experiment. However, whenever the colony counts remain within the historical range of negative and solvent controls such an increase is not considered biologically relevant. - Statistics:
- According to the OECD guideline 471, a statistical analysis of the data is not mandatory.
Results and discussion
Test resultsopen allclose all
- Key result
- Species / strain:
- E. coli WP2 uvr A
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity, but tested up to precipitating concentrations
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- True negative controls validity:
- not examined
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 100
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- True negative controls validity:
- not examined
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- True negative controls validity:
- not examined
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- True negative controls validity:
- not examined
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- True negative controls validity:
- not examined
- Positive controls validity:
- valid
Applicant's summary and conclusion
- Conclusions:
- In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test item did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used.
- Executive summary:
This study was performed to investigate the potential of the test item to induce gene mutations according to the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100, and the Escherichia coli strain WP2 uvrA. The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration, including the controls, was tested in triplicate. The test item was tested at the following concentrations:
Pre-Experiment/Experiment I: 3; 10; 33; 100; 333; 1000; 2500; and 5000 μg/plate
Experiment II:
Strain TA 98: 3, 10; 33; 100; 333; 1000; 2500 and 5000 μg/plate
The remaining strains: 10; 33; 100; 333; 1000; 2500; and 5000 μg/plate
The test item precipitated in the overlay agar in the test tubes from 1000 to 5000 μg/plate in experiment I and from 2500 to 5000 μg/plate in experiment II. Precipitation of the test item in the overlay agar on the incubated agar plates was observed at 5000 μg/plate in both experiments. The undissolved particles had no influence on the data recording. The plates incubated with the test item showed reduced background growth up to 5000 μg/plate in strains TA 1537, TA 98 and TA 100 in experiment I and in strains TA 1535, TA 1537 and TA 98 in experiment II. Toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), were observed in all strains, except of strains TA 1535 and WP2 uvrA in both experiments. No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with the test item at any dose level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance. Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies.
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