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EC number: 200-718-6 | CAS number: 69-89-6
- 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
Endpoint summary
Administrative data
Key value for chemical safety assessment
Genetic toxicity in vitro
Description of key information
In vitro (mutagenic effects - bacterial): OECD Guideline 471. Negative with and without activation. Reliability = 1
Link to relevant study records
- Endpoint:
- in vitro gene mutation study in bacteria
- 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:
- EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- GLP compliance:
- yes
- Type of assay:
- bacterial reverse mutation assay
- Specific details on test material used for the study:
- Purity: > 99%
- 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:
- Type and composition of metabolic activation system:
- source of S9: Molecular Toxicology, Inc.; sourced from male Sprague-Dawley rats induced with phenobarbital and 5,6-benzoflavone.
- method of preparation of S9 mix: freshly prepared on day of use; maintained on ice prior to and during use.
- concentration or volume of S9 mix and S9 in the final culture medium: contained 5% v/v S9 fraction. Sodium phosphate buffer was used as the substitution buffer for plates treated in the absence of S9.
- quality controls of S9: The prepared S9 mix contained the following sterile cofactors: 8 mM MgCl2, 33 mM KCl, 100 mM sodium phosphate buffer pH 7.4, 5 mM glucose-6-phosphate and 4 mM NADP. - Test concentrations with justification for top dose:
- 1.58, 5.0, 15.8, 50, 150, 500, 1580, and 5000 µg/plate. The top dose is the limit recommended by guideline.
- Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: sterile water (vehicle control); DMSO
- Justification for choice of solvent/vehicle: test substance was found to be soluble in sterile water; dilutions in DMSO - Negative solvent / vehicle controls:
- yes
- Remarks:
- vehicle
- Positive control substance:
- other:
- Remarks:
- for all tester strains
- Positive control substance:
- methylmethanesulfonate
- Remarks:
- for tester strain E. coli WP2 uvrA
- Positive control substance:
- other:
- Remarks:
- for tester strain S. typhimurium TA98
- Positive control substance:
- other:
- Remarks:
- for tester strain S. typhimurium TA1537
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- Remarks:
- for tester strain S. typhimurium TA100, TA1535
- Details on test system and experimental conditions:
- NUMBER OF REPLICATIONS:
- Number of cultures per concentration: triplicates
- Number of independent experiments: 2 (Main and Confirmatory)
METHOD OF TREATMENT/ EXPOSURE (MAIN):
- Following materials mixed and added over surface of minimal agar plate: 100 µL of the prepared test substance solutions, negative (vehicle) control, or prepared positive control substance; 500 µL S9 mix or substitution buffer; 100 µL bacteria suspension (ST or EC); and 2000 µL overlay agar maintained at approximately 45°C.
- Plates placed on level surface until agar gelled then inverted and incubated at approximately 37⁰C until growth was adequate for enumeration. Appropriate sterility control check plates (treated with critical components in absence of bacteria) included as a standard procedural check.
TREATMENT AND HARVEST SCHEDULE:
- Exposure duration/duration of treatment: approximately 65 hours
- Harvest after the end of treatment (sampling/recovery times): After incubation, number of colonies per plate counted manually and/or with the aid of a plate counter.
CONFIRMATORY TEST
Employed pre-incubation modification of plate incorporation test. Test or control substances, bacteria suspension, and S9/substitution buffer were incubated under agitation for approximately 30 minutes at approximately 37°C prior to mixing with overlay agar and pouring onto minimal agar plates before proceeding to main test. The study design for the confirmatory test, including strains, dose levels etc. was as described for main test.
METHODS FOR MEASUREMENT OF CYTOTOXICITY
- Method: Evaluated by spectrophotometric optical density measurement. - Evaluation criteria:
- Criteria for Validity: Background lawn for vehicle control plates should appear normal. Mean revertant colony counts for each strain treated with vehicle should lie close to or within expected range taking into account the laboratory historical control range and/or published values. Positive controls (with S9 where required) should produce substantial increases in revertant colony numbers with appropriate bacterial strain. In case where part of study is invalid based on these criteria, detailed results for that part of study will not be reported and affected part of study would be subjected to an automatic repeat at the discretion of the Study Director.
Evaluation of Toxicity: Toxic effects of test substance are indicated by partial or complete absence of background lawn of non-revertant bacteria or a substantial dose-related reduction in revertant colony counts compared with lower dose levels and concurrent vehicle control taking into account laboratory historical control range. Where precipitation obscures observations on condition of background lawn, lawn can be considered normal and intact if revertant colony counts are within expected range based on results for lower dose levels and historical control counts for that strain.
Evaluation of Mutagenicity: For each experimental point, the Mutation Factor (MF) calculated by dividing mean revertant colony count by mean revertant colony count for corresponding concurrent vehicle control. Results considered positive if substantial increase in revertant colony counts. Otherwise, results were considered negative. This increase must be dose related and/or reproducible. Test substance that produces neither a concentration related increase in number of revertant colonies nor a reproducible substantial increase in revertant colonies is considered to be non-mutagenic in test system. - Statistics:
- Calculated means and standard deviations for all quantitative data collected.
- Key result
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- True negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 100
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- True negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- True negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- True negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Key result
- 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
- True negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Additional information on results:
- No signs of toxicity were seen in any of the strains at all dose levels. For all strains, the plates containing 1580 µg and 5000 µg of test substance had precipitate. Contamination was observed in few plates throughout the assay; however, it did not obscure the count with the exception of one replicate in TA98 for the pre-incubation method. Although the one plate was unreadable there were still two replicate plates that were able to be evaluated. For all strains, at least six non-toxic dose levels without precipitate were evaluated, therefore bacterial mutagenicity was adequately assessed. For negative control validity, at least six non-toxic dose levels without precipitate were evaluated, therefore bacterial mutagenicity adequately assessed. For positive control validity, expected substantial increases in revertant colony counts in both absence and presence of S9 in each phase of test confirmed sensitivity of test and activity of S9 mix.
- Conclusions:
- The test substance was negative for mutagenicity in a Bacterial Reverse Mutation Test (Ames Test).
- Executive summary:
The purpose of the study was to evaluate the potential for the test material to induce gene mutations in bacteria using the Ames assay. Point mutations which involve substitution, addition or deletion of one or a few DNA base pairs are detected in amino acid-requiring strains of Salmonella typhimurium (S. typhimurium, ST) and Escherichia coli (E. coli, EC) by their ability to functionally reverse mutations. These reverse mutations result in revertant colonies of bacteria with restored capability to synthesize the essential amino acid. This Ames test was conducted under test guidelines U.S. EPA Health Effects Test Guidelines, OPPTS 870.5100 “Bacterial Reverse Mutation Test” (1998) and OECD Guidelines for Testing of Chemicals, Section 4 (Test No. 471): “Bacterial Reverse Mutation Test" (1997). The test was run with test substance levels of 1.58, 5.0, 15.8, 50, 158, 500, 1580, and 5000 µg/plate, with the high level being the standard limit for the test. The main test was conducted using the plate incorporation method in both the absence and presence of metabolic activation (chemically-induced rat liver S9 mix). The results of the test were confirmed using a similar study design but employing the pre-incubation modification of the Ames test.
The mean revertant colony counts for each strain treated with the vehicle were close to or within the expected range, considering the laboratory historical control range and/or published values. The positive control substances caused the expected substantial increases in revertant colony counts in both the absence and presence of S9 in each phase of the test confirming the sensitivity of the test and the activity of the S9 mix. Therefore, each phase of the test is considered valid. No signs of toxicity were seen in any of the strains at all dose levels. For all strains, the plates containing 1580 µg and 5000 µg of test substance had precipitate. Contamination was observed in few plates throughout the assay; however, it did not obscure the count with the exception of one replicate in TA98 for the pre-incubation method. Although the one plate was unreadable there were still two replicate plates that were able to be evaluated. For all strains, at least six non-toxic dose levels without precipitate were evaluated, therefore bacterial mutagenicity was adequately assessed.There was no concentration-related or substantial test substance related increases in the number of revertant colonies observed with strains TA98, TA100, TA1535, TA1537, or E.coli WP2 uvrA in both the absence and presence of S9 using either the plate incorporation or the pre-incubation method. In conclusion, based on these findings and on the evaluation system used, the test substance did not elicit evidence of bacterial mutagenicity in the Ames assay.
Reference
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Additional information
Justification for classification or non-classification
The test substance was negative for mutagenicity in a Bacterial Reverse Mutation Test (Ames Test). Therefore, the substance does not need to be classified for mutagenicity according to EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.
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