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EC number: 278-817-9 | CAS number: 78014-16-1
- 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
Endpoint summary
Administrative data
Key value for chemical safety assessment
Genetic toxicity in vitro
Description of key information
Ames test: positive in one tester strain (Salmonella th. TA 1535) without metabolic activation
Mouse lymphoma assay: negative with/without metabolic activation
Micronucleus test: negative with/without metabolic activation
Link to relevant study records
- Endpoint:
- in vitro gene mutation study in bacteria
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9 mix (rat)
- Test concentrations with justification for top dose:
- 3, 10, 33, 100, 333, 1000, 2500 and 5000 µg/plate
- Vehicle / solvent:
- deionised water
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- methylmethanesulfonate
- other: 4-nitro-o-phenylene-diamine (4-NOPD), 2-aminoanthracene (2-AA)
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: 1st main experiment: plate incorporation; 2nd main experiment: preincubation
NUMBER OF REPLICATIONS: 3
DETERMINATION OF CYTOTOXICITY
- Method: determination of background lawn - Evaluation criteria:
- A test item is considered as a mutagen if a biologically relevant increase in the number of revertants exceeding the threshold of twice (strains TA 98, TA 100, and WP2 uvrA) or thrice (strains TA 1535 and TA 1537) the colony count of the corresponding solvent control is observed.
A dose dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration.
An increase exceeding the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.
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.
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- without
- Genotoxicity:
- positive
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- with
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium, other: TA 98, TA 100, TA 1537
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- E. coli WP2 uvr A
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Conclusions:
- Positive without metabolic activation in strain TA 1535
Negtaive without metabolic activation in strains TA 1537, TA 98, TA 100 and E. coli WP2 uvrA
Negative with metabolic activation all strains tested
The test item induced gene mutations by base pair changes in the genome of the strain TA 1535 in the absence of metabolic activation. - Executive summary:
This study was performed to investigate the potential of TAT (Triacetonetriamine) to induce gene mutations in the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, and TA 100, and the Escherichia coli strain WP2 uvrA.
The assay was performed with and without liver microsomal activation. Each concentration, including the controls, was tested in triplicate. The test item was tested at the following concentrations:
3; 10; 33; 100; 333; 1000; 2500; and 5000µg/plate
The plates incubated with the test item showed normal background growth up to 5000 Ng/plate with and without metabolic activation in both independent experiments.
No toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), occurred in the test groups with and without metabolic activation.
A substantial and dose dependent increase in revertant colony numbers was observed following treatment with TAT (Triacetonetriamine) in strain TA 1535 in the absence of metabolic activation. The threshold of thrice the number of the corresponding solvent control was exceeded at 2500 Ng/plate. A dose dependent increase in revertant colony numbers was also observed in strain TA 100 without metabolic activation. However, the threshold of twice the number of the corresponding solvent control was not reached.
Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies.
Reference
Endpoint conclusion
- Endpoint conclusion:
- adverse effect observed (positive)
Additional information
Additional information from genetic toxicity in vitro:
According to Regulation (EC) No. 1907/2006, Annex VIII, point 8.4, appropriate in vivo mutagenicity studies shall be considered in case of a positive result in any of the genotoxicity studies in Annex VII or VIII. In the Ames test with triacetonetriamine positive results were obtained in one test strain without metabolic activation. The test item induced gene mutations by base pair changes in the genome of the strain TA 1535 in the absence of metabolic activation. In the available cell mutation assay in mouse lymphoma cells no mutagenic activity with or without metabolic activation was observed. The available in vitro micronucleus test in human lymphocytes also showed clearly negative results.
The registrant has decided to propose an appropriate test in vivo to clarify the genotoxicity observed in vitro. The positive result occured in an in vitro mutagenicity study, therefore the in vivo test needs to focus on mutagenicity as the genotoxic mode of action.
For animals welfare reasons it would be preferable to include the in vivo mutagenicity assessment into another in vivo study which needs to be proposed to fulfil the standard data requirements. The tonnage band of 100 tonnes/year is exceeded. Thus, the performance of a sub-chronic (90 days, OECD TG 408) toxicity study is being proposed according to Annex IX, 8.6.2. The registrant has been evaluated the possibilities to include the in vivo mutagenicity assay into the subchronic toxicity study.
A novel assay, the in vivo PIG-A (endogenous X-linked phosphatidylinositol glycan, Class A, Pig-a in rodents and PIG-A in humans) gene mutation assay, shows promise for regulatory applications as a reporter of in vivo mutation, by integration of gene mutation measurement into repeat-dose toxicology studies. [1, 7] The measurement of PIG-A mutants by counting cells with the GPI-negative (glycosylphosphatidyl inositol) phenotype has proved to be effective to measure mutant frequency in peripheral blood cells of humans and of others animals. [2] Currently the PIG-A assay has been extended for its use in human erythrocytes. Its advantages are the applicability across species, its simplicity and statistical power, and the relatively non-invasive nature. [3] A collaborative international trial was conducted to evaluate the reproducibility and transferability of an in vivo mutation assay based on the enumeration of CD59-negative rat erythrocytes across 14 laboratories. The methodology was demonstrated to be reproducible and a good transferability was evident from the similar kinetics and magnitude of the dose-related responses that were observed among different laboratories. The results of the trial demonstrate that the method is a robust in vivo mutation assay that is readily transferable across laboratories. [4] The erythrocyte-based Pig-a mutation assay was successfully used to assess the ability of to discriminate between genotoxic and non-genotoxic modes of action, for methyl carbamate (MC) and ethyl carbamate (EC). [8] It has been demonstrated recently that the utility and sensitivity of the Pig-a in vivo gene mutation assay, can be easily integrated, along with other standard genotoxicity endpoints, into e.g. 28-day rodent toxicity studies. [5] It has been concluded that the PIG-A assay could be a useful and sensitive endpoint for a repeat dose protocol and complements other genotoxicity endpoints. [6]
A committee entitled “Relevance and Follow-up of Positive Results from In Vivo Genetic Toxicity Testing” (IVGT), initiated by the Health and Environmental Sciences Institute (HESI), recommends the Pig-a assay (in combination with the micronucleus assay (MN)) in repeat-dose toxicity studies “as it would allow for the first time the simple assessment of aneugenicity, clastogenicity, and mutagenicity” [9]
The US EPA has concluded that although the Pig-A gene mutation assay does not have an OECD Test guideline yet, it is a promising new in vivo mutation test. US EPA encourage the incorporation of genotoxicity endpoints into routine toxicology studies where scientifically feasible. [http://www.epa.gov/pesticide-science-and-assessing-pesticide-risks/advances-genetic-toxicology-and-integration-vivo]
Taking into account the relatively simple integration of the PIG-A assay into repeated-dose toxicity studies, the low volume of blood needed, the reduction of animal use and the fact that no expensive transgenic animals are required, the consideration of the PIG-A assay being a suitable methodology for the in vivo verification of a mutagenic potential observed in vitro is reasonable.
Therefore the registrant proposes to perform a PIG-A assay as an integral part of the also proposed subchronic oral repeated-dose toxicity study (OECD 408) according to Annex IX, 8.6.2, in order to verify the positive results obtained in the Ames test and fulfilling the data requirements according to Regulation (EC) No. 1907/2006, Annex VIII, point 8.4.
[1] Dobrovolsky VN et al., The in vivo Pig-a gene mutation assay, a potential tool for regulatory safety assessment, Environ Mol Mutagen. 2010 Oct-Dec;51(8-9):825-35,
[2] Peruzzi et al., The use of PIG-A as a sentinel gene for the study of the somatic mutation rate and of mutagenic agents in vivo, Mutat Res. 2010 Jul-Sep;705(1):3-10,
[3] Dertinger SD et al., Human erythrocyte PIG-A assay: an easily monitored index of gene mutation requiring low volume blood samples, Environ Mol Mutagen. 2015 May;56(4):366-77,
[4] Dertinger SD et al., International Pig-a gene mutation assay trial: evaluation of transferability across 14 laboratories, Environ Mol Mutagen. 2011 Dec;52(9):690-8,
[5] Stankowski LF et al., Integration of Pig-a, micronucleus, chromosome aberration and comet assay endpoints in a 28-day rodent toxicity study with urethane, Mutagenesis. 2015 May;30(3):335-42,
[6] Gunther WC et al., Evaluation of the Pig-a, micronucleus, and comet assay endpoints in a 28-day study with ethyl methanesulfonate, Environ Mol Mutagen. 2014 Jul;55(6):492-9,
[7] Bhalli JA et al., Sensitivity of the Pig-a assay for detecting gene mutation in rats exposed acutely to strong clastogens, Mutagenesis (2013),
[8] Bemis JC et al., Rat Pig-a mutation assay responds to the genotoxic carcinogen ethyl carbamate but not the non-genotoxic carcinogen methyl carbamate, Mutagenesis (2015)
[9] Schuler M et al., Need and potential value of the Pig-a in vivo mutation assay – a HESI perspective, Environ. Mol. Mutagen., 52 (2011), pp. 685–689
Justification for classification or non-classification
Based on the currently available experimental in vitro data, no conclusion on the in vivo relevance of the positive results obtained in the Ames test can be made. Re-evaluation of the classification will be performed after in vivo data has become available.
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
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