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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
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:
- 010-01-12 - 2010-01-29
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 010
- Report date:
- 2010
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
Test material
- Reference substance name:
- Recombinant spider silk protein eADF4(C16) derived from E. Coli by fermentation
- Molecular formula:
- not applicable, UVCB
- IUPAC Name:
- Recombinant spider silk protein eADF4(C16) derived from E. Coli by fermentation
- Test material form:
- liquid
Constituent 1
Method
- Target gene:
- uvrB gene
Species / strain
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
- Metabolic activation:
- with and without
- Metabolic activation system:
- Mammalian Microsomal Fraction S9 Mix
- Test concentrations with justification for top dose:
- 0.0316, 0.100, 0.316, 1.0, 2.5 and 5.0 µl./plate
- Vehicle / solvent:
- 0.9% NaCl
Controls
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- methylmethanesulfonate
- other: 2-aminoanthracene, 4-nitro-o-phenylene-diamine
- Details on test system and experimental conditions:
- Pre-Experiment for Toxicity
The toxicity of the lest item was determined with tester strains TA 98 and TA 100 in a pre-experiment. Eight concentrations were tested for toxicity and induction of mutations with three plates each. The experimental conditions in this pre¬experiment were the same as described below for the main experiment 1 (plate incorporation test).
Toxicity may be delected by a clearing or rather diminution of the background lawn or a reduction in the number of revertants down to a mutation factor of approximately < 0.5 in relation to the solvent control.
The test item was tested in the pre-experirnent with the following concentrations:
0.00316, 0.0100, 0.0316, 0.100, 0.316, 1.0. 2.5 and 5.0 µL/plate
Exposure Concentrations
The lest item concentrations to be applied in the main experiments were chosen according to the results of the pre-experiment. 5.0 µL/plate was selected as the maximum concentration. The concentration range covered two logarithmic decades. Two independent experiments were performed with the following concentrations:
0.0316, 0.100, 0.316, 1.0, 2.5 and 5.0 µL/plate
Experimental Performance
For the plate incorporation method the following materials were mixed in a test tube and poured over the surface of a minimal agar plate:
100 µl: Test solution at each dose level, solvent control, negative control or reference mutagen solution (positive control).
500 µL: S9 mix (for testing with metabolic activation) or S9 mix substitution buffer (for testing without metabolic activation).
100 µL: Bacteria suspension (cf. Preparation of Bacteria, pre¬culture of the strain).
2000 µL: Overlay agar.
For the pre-incubation method 100µ L of the test item preparation was pre¬incubated with the tester strains (100 pL) and sterile buffer or the metabolic activation system (500 µL) for 60 minutes al 37 °C prior Io adding the overlay agar (2000µL) and pouring onto the surface of a minimal agar plate.
For each strain and dose level, including the controls, three plates were used.
After solidification the plates were inverted and incubated at 37 °C for at least 48 h in the dark.
Data Recording
The colonies were counted using a ProtoCOL counter. If precipitation of the test item precluded automatic counting the revertant colonies were counted by hand. In addition, tester strains with a low spontaneous mutation frequency like TA 1535 and TA 1537 were counted manually. - Evaluation criteria:
- A test item is considered as mutagenic if:
- a clear and dose-related increase in the number of revertants occurs and/or
- a biologically relevant positive response for at least one of the dose groups occurs
in al least one tester strain with or without metabolic activation.
A biologically relevant increase is described as follows:
- if in tester strains TA 100 and TA 102 the number of reversions is al least twice as high
- if in tester strains TA 98. TA 1535 and TA 1537 the number of reversions is at least three times higher than the reversion rate of the solvent control.
Results and discussion
Test resultsopen allclose all
- Key result
- Species / strain:
- S. typhimurium TA 102
- 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
- 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
- Untreated negative controls validity:
- valid
- 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
- Untreated negative controls validity:
- valid
- 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
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- 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
- Untreated negative controls validity:
- valid
- 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, eADF4(C16) did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used.
Therefore, eADF4(C16) is considered to be non-mutagenic in this bacterial reverse mutation assay.
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