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EC number: - | CAS number: -
- 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
Under the present test conditions the test substance was tested up to a cytotoxic concentration of 316 µg/plate caused no mutagenic effect in the Salmonella typhimurium strains TA98, TA100, TA102, TA1535 and TA1537 neither in the plate incorporation test nor in the preincubation test each carried out without and with metabolic activation (LPT 2013).
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
- Study period:
- 2012-09-19 to 2013-01-11
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Guideline study; GLP study without deviations
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Version / remarks:
- 1997
- Deviations:
- no
- 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:
- EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
- Version / remarks:
- OPPTS changed its name to OCSPP,
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
- Target gene:
- mutated gene loci resposible for histidine auxotropy
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
- Details on mammalian cell type (if applicable):
- - obtained from Trinova Biochem according to Dr. Bruce N. AMES,
- Additional strain / cell type characteristics:
- other: histidine auxotroph
- Metabolic activation:
- with and without
- Metabolic activation system:
- Arochlor 1254 induced rat liver S9; male rats
- Test concentrations with justification for top dose:
- Plate incorporation test: 1.0, 3.16, 10.0, 31.6, 100 and 316 µg per plate;
Preincubation test: 1.0, 3.16, 10.0, 31.6, 100 and 316 µg per plate - Vehicle / solvent:
- 50 mg of test item were completely dissolved in 10 mL aqua ad iniectabilia by using an ultrasonic bath (45 kHz) at 37°C for 60 minutes to a
concentration of 5 mg/mL. In the preliminary test the administration volume had to be increased from the generally employed 100 μL to
1000 μL per plate to achieve a concentration of 5000 μg/plate. In the main study an administration volume of 100 μL/plate was used for all
concentrations. The vehicle served as the negative control. Fresh preparations of the test item were used for the treatment in all experimental parts. - Untreated negative controls:
- no
- Remarks:
- solvent test will be used as negative reference item
- Negative solvent / vehicle controls:
- yes
- Remarks:
- aqua ad iniectabilia
- True negative controls:
- no
- Positive controls:
- yes
- Remarks:
- for details see below
- Positive control substance:
- other: without metabolic activation: sodium azide in aqua ad iniectabilia for TA 1535 and TA 100, 2-nitroflurene in DMSO for TA 98, 9-amino-acridine in ethanol abs. for TA 1537, Mitomycin C in DMSO for TA 102
- Remarks:
- with metabolic activation: 2-aminoanthracene in DMSO for TA 100, TA 1535, Benzo(a)pyrene in DMSO for TA 98, TA 102, TA 1537
- Details on test system and experimental conditions:
- Bacterial Reverse Mutation Test
SYSTEM OF TESTING
- Pre-Experiment: plate incorporation cytotoxicity test (+/- metabolic activation) with strain TA 100,
The test item was examined in two preliminary cytotoxicity tests (plate incorporation test without and with metabolic activation) in test strain
TA100. Ten concentrations ranging from 0.316 to 5000 µg test item/plate were tested. Cytotoxicity (scarce background lawn and reduction of the number of revertants by more than 50%) was noted starting at a concentration of 316 μg/plate.
Hence, 316 μg test item/plate were chosen as top concentration for the main study in the plate incorporation test and in the preincubation test.
- Main test: 1st - Standard plate incorporation method, 2nd - Preincubation method
- Metabolic activation assay: Arochlor 1254 induced rat liver S9 fraction, the protein content of the S9 fraction was 33.1 mg/mL S9, cytochrome
P-450: 0.40 nmol/mg protein
ADMINISTRATION
- Dosing:
* Plate incorporation test: 1.0, 3.16, 10.0, 31.6, 100 and 316 µg per plate
* Preincubation test: 1.0, 3.16, 10.0, 31.6, 100 and 316 µg per plate
- Data : 2 independent experiments with and without metabolic activation
- Number of replicates: 3 per concentration and experiment
- Positive and negative control groups and treatment:
- without metabolic activation:
* sodium azide in aqua ad iniectabilia for TA 1535 and TA 100, 10 µg/plate
* 2-nitroflurene in DMSO for TA 98, 10 µg/plate
* 9-amino-acridine in ethanol abs. for TA 1537, 100 µg/plate
* Mitomycin C in DMSO for TA 102, 10 µg/plate
- with metabolic acivation
* 2-aminoanthracene in DMSO for TA 100 and TA 1535, 2 µg/plate
* Benzo(a)pyrene in DMSO for TA 98, TA 102 and 1537, 10 µg/plate
- negative control: the vehicle aqua ad iniectabilia was used as negative reference item (all test strains).
- Incubation time: 48 h to 72 h at 37 °C in the dark
- Pre-incubation time: 20 min at 37 °C;
NUMBER OF REPLICATIONS: 3 per concentration and experiment
NUMBER OF CELLS EVALUATED: approximately 10E8 viable cells in the late exponential or early stationary phase
DETERMINATION OF CYTOTOXICITY
- Method: In the plate incorporation test and in the preincubation test, each carried out without and with metabolic activation cytotoxicity (scarce
background lawn and reduction of the number of revertants) was noted at the top concentration of 316 μg/plate.
- Evaluation criteria:
- The statistical evaluation of the results of the AMES test is still under discussion. In our laboratory, a test item is considered to show a
positive response if
- at one or more concentrations the number of revertants is reproducibly increased in at least one strain with or without metabolic activation. A
2-fold increase in comparison to the solvent control is regarded as being relevant for a positive response in the strains TA98, TA100 and TA102. For the strains TA1535 and TA1537 a 3-fold increase represents a biological relevant effect. The Mann and Whitney test (p ≤ 0.05, see section 6,
reference 3.) may be used to determine statistical significance.
Or
- a concentration-related increase over the range tested in the number of the revertants per plate is observed. The Spearman's rank correlation
coefficient (section 6, reference 3.) may be applied.
Biological relevance of the results should be considered first.
Positive results have to be reproducible and the histidine independence of the revertants has to be confirmed by streaking random samples on
histidine-free agar plates.
A test item for which the results do not meet the above mentioned criteria is considered as non-mutagenic in the AMES test. - Statistics:
- According to the OECD Guideline 471, a statistical analysis of the data is not mandatory
- Key result
- Species / strain:
- S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- was noted at the top concentration of 316 μg/plate, in the plate incorporation test and in the preincubation test, each carried out without and with metabolic activation
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not examined
- Positive controls validity:
- valid
- Additional information on results:
- GENTOXIC EFFECTS:
- With metabolic activation: negativ
- Without metabolic activation: negativ
CYTOTOXICITY EFFECTS:
- plate incorporation test without and with metabolic activation: was noted at the top concentration of 316 μg/plate
- preincubation test without metabolic activation: was noted at the top concentration of 316 μg/plate - Remarks on result:
- other: all strains/cell types tested
- Remarks:
- Migrated from field 'Test system'.
- Conclusions:
- Interpretation of results (migrated information):
negative
Under the present test conditions the test item tested up to a cytotoxic concentration of 316 µg/plate caused no mutagenic effect in the Salmonella
typhimurium strains TA98, TA100, TA102, TA1535 and TA1537 neither in the plate incorporation test nor in the preincubation test each carried out without and with metabolic activation. - Executive summary:
The purpose of this study was to evaluate the test substance for mutagenic activity (gene mutation) in bacteria without and with the addition of a mammalian metabolic activation system as originally described by AMES et al. (1973, 1975) and revised by MARON and (1983).
The test item was examined in the 5 Salmonella typhimurium strains TA98, TA100, TA102, TA1535 and TA1537 in two independent experiments, each carried out without and with metabolic activation (a microsomal preparation derived from Aroclor 1254-induced rat liver). The first experiment was carried out as a plate incorporation test and the second as a preincubation test.
Test item was completely dissolved in aqua ad iniectabilia.The vehicle served as the negative control.
Preliminary test
The test item was examined in two preliminary cytotoxicity tests (plate incorporation test without and with metabolic activation) in test strain TA100. Ten concentrations ranging from 0.316 to 5000 µg test item/plate were tested.Cytotoxicity (scarce background lawn and reduction of the number of revertants by more than 50%) was noted startingat a concentration of 316 µg/plate.
Hence, 316 µg test item/plate were chosen as top concentration for the main study in the plate incorporation test and in the preincubation test.
Main study
Six concentrations ranging from 1.0 to 316 µg test item/plate were employed in the plate incorporation test and in the preincubation test, each carried out without and with metabolic activation.
Cytotoxicity
In the plate incorporation test and in the preincubation test, each carried out without and with metabolic activation cytotoxicity (scarce background lawn and reduction of the number of revertants) was notedat the top concentration of 316 µg/plate.
Mutagenicity
No increase in revertant colony numbers as compared with control counts was observed for the test item, tested up to a cytotoxic concentration of 316 µg/plate, in any of the 5 test strains in two independent experiments without and with metabolic activation, respectively (plate incorporation and preincubation test).
The positive control items showed a significant increase in the number of revertant colonies of the respective test strain and confirmed the validity of the test conditions and the sensitivity of the test system.
In conclusion, under the present test conditions the test item tested up to a cytotoxic concentration of 316 µg/plate caused no mutagenic effect in the Salmonella typhimurium strains TA98, TA100, TA102, TA1535 and TA1537 neither in the plate incorporation test nor in the preincubation test each carried out without and with metabolic activation.
Reference
see attchached document
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Additional information
Under the present test conditions Indate (2-), hexachlorododeca-µ-methoxy-µ6-oxohexa-, hydrogen, compd. with N-methylmethanamine (1:2:2) caused no mutagenic effect in the Salmonella typhimurium strains TA98, TA100, TA102, TA1535 and TA1537 neither in the plate incorporation test nor in the preincubation test each carried out without and with metabolic activation.
Justification for selection of genetic toxicity endpoint
Annex VII requires an in vitro gene mutation assay in bacteria. Therefore the Salmonella reverse mutation assay has been chosen.
Justification for classification or non-classification
According to the EC Regulation 1272/2008 and subsequent regulations on classification, labelling and packaging of substances and mixtures, the test item is not genotoxic and has not to be classified.
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