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EC number: 277-761-2 | CAS number: 74196-18-2
- 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:
- other: read-across from supporting substance (structural analogue or surrogate)
- Adequacy of study:
- key study
- Study period:
- From 23. Oct. to 03. Dec. 2014
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- The complete read-across justification is detailed in section 13; source study has reliability 1.
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 015
- Report date:
- 2015
Materials and methods
Test guidelineopen allclose all
- 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
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
Test material
- Reference substance name:
- Similar Substance 01
- IUPAC Name:
- Similar Substance 01
- Test material form:
- solid
Constituent 1
Method
- Target gene:
- HIS /TRP
Species / strainopen allclose all
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
- Additional strain / cell type characteristics:
- not applicable
- Species / strain / cell type:
- E. coli WP2 uvr A
- Additional strain / cell type characteristics:
- not applicable
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9-Mix (rat and hamster)
- Test concentrations with justification for top dose:
- - 33, 100, 333, 1000, 2600 and 5200 μg/plate (SPT)
- 33, 100, 333, 1000, 2600 and 5200 μg/plate (Prival test) - Vehicle / solvent:
- - Vehicle/solvent used: DMSO
Controlsopen allclose all
- Untreated negative controls:
- yes
- Remarks:
- treated with soft agar, S9 mix, buffer, vehicle or the test item without the addition of tester strains
- Negative solvent / vehicle controls:
- yes
- Remarks:
- contains vehicle used at the same concentration and volume (all strains)
- Positive controls:
- yes
- Positive control substance:
- other: 2-aminoanthracene
- Remarks:
- With rat liver S9 mix
- Untreated negative controls:
- yes
- Remarks:
- treated with soft agar, S9 mix, buffer, vehicle or the test item without the addition of tester strains
- Negative solvent / vehicle controls:
- yes
- Remarks:
- contains vehicle used at the same concentration and volume (all strains)
- Positive controls:
- yes
- Positive control substance:
- other: 2-aminoanthracene
- Remarks:
- With hamster liver S9 mix
- Untreated negative controls:
- yes
- Remarks:
- treated with soft agar, S9 mix, buffer, vehicle or the test item without the addition of tester strains
- Negative solvent / vehicle controls:
- yes
- Remarks:
- contains vehicle used at the same concentration and volume (all strains)
- Positive controls:
- yes
- Positive control substance:
- 9-aminoacridine
- other:
- not specified
- Remarks:
- Without S9 mix
- Details on test system and experimental conditions:
- STANDARD PLATE TEST
The experimental procedure of the standard plate test (plate incorporation method) was based on the method of Ames et al.
Salmonella typhimurium
Test tubes containing 2 ml portions of soft agar (overlay agar), which consists of 100 ml agar (0.8 % w/v agar + 0.6% w/v NaCl) and 10 ml amino acid solution (minimal amino acid solution for the determination of mutants: 0.5 mM histidine + 0.5 mM biotin) were kept in a water bath at about 42 - 45 °C, and the remaining components were added in the following order:
- 0.1 ml test solution or vehicle (negative control)
- 0.1 ml fresh bacterial culture
- 0.5 ml S9 mix (with metabolic activation) or 0.5 ml phosphate buffer (without metabolic activation)
After mixing, the samples were poured onto Minimal glucose agar plates (Moltox Molecular Toxicology, Inc.; Boone, NC 28607; USA) within approx. 30 seconds and incubated at 37 °C for 48 to 72 hours in the dark, and the bacterial colonies (his+ revertants) were counted using the Sorcerer Image Analysis System with the software program Ames Study Manager (Perceptive Instruments Ltd., Haverhill, UK). Colonies were counted manually, if precipitation of the test substance hinders the counting using the Image Analysis System.
Escherichia coli
Test tubes containing 2 ml portions of soft agar (overlay agar), which consists of 100 ml agar (0.8 % w/v agar + 0.6% w/v NaCl) and 10 ml amino acid solution (minimal amino acid solution for the determination of mutants: 0.5 mM tryptophan) were kept in a water bath at about 42 - 45°C, and the remaining components were added in the following order:
- 0.1 ml test solution or vehicle (negative control)
- 0.1 ml fresh bacterial culture
- 0.5 ml S9 mix (with metabolic activation) or 0.5 ml phosphate buffer (without metabolic activation)
After mixing, the samples were poured onto Minimal glucose agar plates (Moltox Molecular Toxicology, Inc.; Boone, NC 28607; USA) within approx. 30 seconds. After incubation at 37 °C for 48 to 72 hours in the dark, the bacterial colonies (his+ revertants) were counted using the Sorcerer Image Analysis System with the software program Ames Study Manager (Perceptive Instruments Ltd., Haverhill, UK). Colonies were counted manually, if precipitation of the test substance hinders the counting using the Image Analysis System.
PRIVAL PREINCUBATION TEST
This experimental procedure is based on the method described by Yahagi et al. and Matsushima et al. and has been modified further to include reductive conditions by Prival et al.
0.1 ml test solution or vehicle (negative control), 0.1 ml bacterial suspension and 0.5 ml S9 mix (with metabolic activation) or phosphate buffer (without metabolic activation) were incubated at 30 °C for 30 minutes using a shaker. Subsequently, 2 ml soft agar which consists of 100 ml agar (0.8% w/v agar + 0.6% w/v NaCl) and 10 ml amino acid solution (minimal amino acid solution for the determination of mutants: 0.5 mM histidine + 0.5 mM biotin or 0.5 mM tryptophan) were added. After mixing, the samples were poured onto the Vogel-Bonner agar plates (minimal glucose agar plates) within approx. 30 seconds. Composition of the minimal glucose agar:
- 980 ml purified water
- 20 ml Vogel-Bonner E medium
- 15 g Difco bacto agar
- 5 g D-glucose, monohydrate
After incubation at 37 °C for 48 to 72 hours in the dark, the bacterial colonies were counted. The colonies were counted using the Sorcerer Image Analysis System with the software program Ames Study Manager (Perseptive Instruments Ltd., Haverhill, UK). Colonies were counted manually, if precipitation of the test substance hindered the counting using the Image Analysis System.
DETERMINATION OF CYTOTOXICITY
- Method: relative total growth - Evaluation criteria:
- Acceptance criteria
Generally, the experiment was considered valid if the following criteria were met:
- the number of revertant colonies in the negative controls was within the range of the historical negative control data for each tester strain;
- the sterility controls revealed no indication of bacterial contamination;
- the positive control substances both with and without S9 mix induced a distinct increase in the number of revertant colonies within the range of the historical positive control data or above;
- fresh bacterial culture containing approximately 10^9 cells per ml were used.
Assessment criteria
The test item was considered positive in this assay if the following criteria were met: a dose-related and reproducible increase in the number of revertant colonies, i.e. at least doubling (bacteria strains with high spontaneous mutation rate, like TA 98, TA 100 and E.coli WP2 uvrA) or tripling (bacteria strains with low spontaneous mutation rate, like TA 1535 and TA 1537) of the spontaneous mutation rate in at least one tester strain either without S9 mix or after adding a metabolizing system.
A test item was generally considered non-mutagenic in this test if: the number of revertants for all tester strains were within the historical negative control range under all experimental conditions in at least two experiments carried out independently of each other.
Results and discussion
Test resultsopen allclose all
- Species / strain:
- S. typhimurium, other: TA 1535, TA 1537, TA 98, TA 100
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- 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:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- TOXICITY
A bacteriotoxic effect (decrease in the number of his+ or trp+ revertants) was occasionally observed in the standard plate test depending on the strain and test conditions from about 1000 μg/plate onward.
In the prival preincubation assay bacteriotoxicity (reduced his- background growth, decrease in the number of his+ or trp+ revertants) was occasionally observed depending on the strain and test conditions from about 1000 μg/plate onward.
SOLUBILITY
Test substance precipitation was found from about 2600 μg/plate onward with and without S9 mix.
CONTROL
The number of revertant colonies in the negative controls was within or marginally below the range of the historical negative control data for each tester strain, with or without metabolic activation.
The positive control substances both with and without S9 mix induced a significant increase in the number of revertant colonies within the range of the historical positive control data or above.
Any other information on results incl. tables
Table 1: Results of the standard plate test without metabolic activation.
Strain | Test group | Dose (µg/plate) | Mean revertants per plate | SD | Factor | Individual revertant colony counts |
TA 1535 | DMSO | - | 14.7 | 3.1 | - | 14, 12, 18 |
test item | 33 | 11.7 | 1.5 | 0.8 | 13, 12, 10 | |
100 | 11.3 | 6.0 | 0.8 | 17, 12, 5 | ||
333 | 12.7 | 5.8 | 0.9 | 16, 16, 6 | ||
1000 | 6.7 | 2.1 | 0.5 | 9, 6, 5 | ||
2600 | 6.7 | 4.0 | 0.5 | 3 P, 6 P, 11 P | ||
5200 | 5.0 | 3.0 | 0.3 | 2 P, 5 P, 8 P | ||
MNNG | 5.0 | 4180.7 | 120.7 | 285 | 4049, 4286, 4207 | |
TA 100 | DMSO | - | 45 | 6.2 | - | 38, 50, 47 |
test item | 33 | 57.3 | 5.0 | 1.3 | 58, 62, 52 | |
100 | 61.0 | 5.2 | 1.4 | 58, 58, 67 | ||
333 | 53.3 | 13.0 | 1.2 | 66, 40, 54 | ||
1000 | 51.7 | 4.0 | 1.1 | 48, 51, 56 | ||
2600 | 53.0 | 7.8 | 1.2 | 48 P, 62 P, 49 P | ||
5200 | 45.0 | 7.0 | 1.0 | 42 P, 40 P, 53 P | ||
MNNG | 5.0 | 3017.7 | 134 | 67.1 | 2999, 2894, 3160 | |
TA 1537 | DMSO | - | 8.0 | 1.7 | - | 10, 7, 7 |
test item | 33 | 7.3 | 1.5 | 0.9 | 6, 7, 9 | |
100 | 8.0 | 5.0 | 1.0 | 13, 3, 8 | ||
333 | 7.0 | 3.5 | 0.9 | 3, 9, 9 | ||
1000 | 6.3 | 2.3 | 0.8 | 5, 5, 9 | ||
2600 | 7.7 | 2.9 | 1.0 | 6 P, 6 P, 11 P | ||
5200 | 6.7 | 4.7 | 0.8 | 3 P, 12 P, 5 P | ||
AAC | 100.0 | 794.3 | 177.1 | 99.3 | 859, 930, 594 | |
TA 98 | DMSO | - | 27.0 | 2.6 | - | 30, 26, 25 |
test item | 33 | 27.7 | 3.8 | 1.0 | 25, 32, 26 | |
100 | 24.7 | 4.0 | 0.9 | 27, 20, 27 | ||
333 | 25.7 | 4.0 | 1.0 | 30, 25, 22 | ||
1000 | 19.7 | 1.5 | 0.7 | 20, 18, 21 | ||
2600 | 13.3 | 4.9 | 0.5 | 11 P, 10 P, 19 P | ||
5200 | 10.3 | 5.7 | 0.4 | 12 P, 15 P, 4 P | ||
NOPD | 10.0 | 360.3 | 31.0 | 13.3 | 373, 383, 325 | |
E. coli | DMSO | - | 48.7 | 9.8 | - | 43, 43, 60 |
test item | 33 | 50.0 | 4.6 | 1.0 | 45, 51, 54 | |
100 | 58.0 | 10.4 | 1.2 | 52, 52, 70 | ||
333 | 58.7 | 4.9 | 1.2 | 62, 53, 61 | ||
1000 | 46.3 | 5.9 | 1.0 | 42, 53, 44 | ||
2600 | 45.0 | 19.0 | 0.9 | 40 P, 66 P, 29 P | ||
5200 | 35.3 | 12.4 | 0.7 | 21 P, 42 P, 43 P | ||
4-NQO | 5.0 | 1030.3 | 5.1 | 21.2 | 1026, 1029, 1036 |
DMSO = dimethyl sulfoxide
MMNG = N-methyl-N'-nitro-N-nitrosoguanidine: 5 µg/plate in DMSO
AAC = 9-aminoacradine 100 µg/plate in DMSO
NOPD = 4-nitro-o-phenylenediamine: 10 µg/plate in DMSO
4-NQO = 4-nitroquinoline-N-oxide 5 μg/plate in DMSO
SD = standard deviation
Table 2: Results of the standard plate test with metabolic activation.
Strain | Test group | Dose (µg/plate) | Mean revertants per plate | SD | Factor | Individual revertant colony counts |
TA 1535 | DMSO | - | 9.7 | 2.1 | - | 9, 12, 8 |
test item | 33 | 12.0 | 1.7 | 1.2 | 11, 14, 11 | |
100 | 10.7 | 0.6 | 1.1 | 11, 11, 10 | ||
333 | 9.7 | 3.2 | 1.0 | 12, 11, 6 | ||
1000 | 8.7 | 4.0 | 0.9 | 11, 4, 11 | ||
2600 | 8.0 | 1.0 | 0.8 | 9 P, 7 P, 8 P | ||
5200 | 3.7 | 2.1 | 0.4 | 6 P, 3 P, 2 P | ||
2-AA | 2.5 | 188.0 | 21 | 19.4 | 167, 209, 188 | |
TA 100 | DMSO | - | 57.0 | 7.0 | - | 49, 62, 60 |
test item | 33 | 58.7 | 5.9 | 1.0 | 61, 63, 52 | |
100 | 69.3 | 8.0 | 1.2 | 77, 61, 70 | ||
333 | 63.0 | 5.2 | 1.1 | 69, 60, 60 | ||
1000 | 43.7 | 5.1 | 0.8 | 48, 38, 45 | ||
2600 | 55.3 | 2.9 | 1.0 | 57 P, 52 P, 57 P | ||
5200 | 50.3 | 2.1 | 0.9 | 51 P, 48 P, 52 P | ||
2-AA | 2.5 | 1815.0 | 420.0 | 31.8 | 2084, 2030, 1331 | |
TA 1537 | DMSO | - | 8.0 | 3.5 | - | 10, 10, 4 |
test item | 33 | 7.0 | 1 | 0.9 | 6, 7, 8 | |
100 | 5.3 | 1.2 | 0.7 | 6, 6, 4 | ||
333 | 6.0 | 1.7 | 0.8 | 4, 7, 7 | ||
1000 | 7.3 | 2.9 | 0.9 | 9, 4, 9 | ||
2600 | 5.3 | 1.5 | 0.7 | 7 P, 4 P, 5 P | ||
5200 | 6.0 | 1.0 | 0.8 | 5 P, 6 P, 7 P | ||
2-AA | 2.5 | 154.7 | 32.3 | 19.3 | 192, 137, 135 | |
TA 98 | DMSO | - | 26.3 | 2.3 | - | 25, 29, 25 |
test item | 33 | 24.0 | 6.6 | 0.9 | 31, 23, 18 | |
100 | 23.3 | 2.3 | 0.9 | 22, 22, 26 | ||
333 | 20.3 | 3.1 | 0.8 | 23, 17, 21 | ||
1000 | 21.7 | 4.5 | 0.8 | 20, 18, 21 | ||
2600 | 17.0 | 10.0 | 0.6 | 27 P, 7 P, 17 P | ||
5200 | 12.7 | 5.9 | 0.5 | 6 P, 17 P, 15 P | ||
2-AA | 2.5 | 1577.3 | 252.4 | 59.9 | 1286, 1716, 1730 | |
E. coli | DMSO | - | 59.7 | 11.4 | - | 47, 63, 69 |
test item | 33 | 57.3 | 4.7 | 1.0 | 61, 52, 59 | |
100 | 55.3 | 9.1 | 0.9 | 47, 65, 54 | ||
333 | 56.0 | 1.7 | 0.9 | 58, 55, 55 | ||
1000 | 54.0 | 2.6 | 0.9 | 56, 51, 55 | ||
2600 | 51.7 | 1.5 | 0.9 | 50 P, 53 P, 52 P | ||
5200 | 33.7 | 2.1 | 0.6 | 33 P, 32 P, 36 P | ||
2-AA | 60.0 | 228.0 | 37.5 | 3.8 | 265, 190, 229 |
DMSO = dimethyl sulfoxide
SD = standard devation
2-AA = 2-aminoanthracene
Table 3: Results of the Prival preincubation test without metabolic activation.
Strain | Test group | Dose (µg/plate) | Mean revertants per plate | SD | Factor | Individual revertant colony counts |
TA 1535 | DMSO | - | 13.3 | 2.1 | - | 15, 14, 11 |
test item | 33 | 15.7 | 4.2 | 1.2 | 19, 17, 11 | |
100 | 10.3 | 4.0 | 0.8 | 14, 11, 6 | ||
333 | 12.7 | 2.5 | 1.0 | 13, 15, 10 | ||
1000 | 11.7 | 0.6 | 0.9 | 12, 11, 12 | ||
2600 | 5.3 | 2.3 | 0.4 | 8 P, 4 P, 4 P | ||
5200 | 8.3 | 3.1 | 0.6 | 11 P, 5 P, 9 P | ||
MNNG | 5.0 | 2292.3 | 284.0 | 171.9 | 2066, 2200, 2611 | |
TA 100 | DMSO | - | 32.0 | 4.0 | - | 32, 36, 28 |
test item | 33 | 30.3 | 4.5 | 0.9 | 35, 30, 26 | |
100 | 32.0 | 3.5 | 1.0 | 34, 34, 28 | ||
333 | 24.7 | 3.1 | 0.8 | 24, 28, 22 | ||
1000 | 13.3 | 4.0 | 0.4 | 17, 9, 14 | ||
2600 | 25.3 | 2.3 | 0.8 | 24 P, 28 P, 24 P | ||
5200 | 17.3 | 2.5 | 0.5 | 17 P, 15 P, 20 P | ||
MNNG | 5.0 | 1520.3 | 241.5 | 47.5 | 1371, 1391, 1799 | |
TA 1537 | DMSO | - | 7.0 | 1.0 | - | 8, 6, 7 |
test item | 33 | 7.0 | 2.6 | 1.0 | 4, 9, 8 | |
100 | 3.7 | 0.6 | 0.5 | 3, 4, 4 | ||
333 | 6.0 | 1.0 | 0.9 | 7, 5, 6 | ||
1000 | 5.3 | 1.5 | 0.8 | 4, 5, 7 | ||
2600 | 3.0 | 1.0 | 0.4 | 3 P, 2 P, 4 P | ||
5200 | 4.3 | 2.5 | 0.6 | 7 P, 4 P, 2 P | ||
AAC | 100 | 1010.0 | 312.7 | 144.3 | 1223, 651, 1156 | |
TA 98 | DMSO | - | 28.0 | 1.0 | - | 27, 29, 28 |
test item | 33 | 14.3 | 4.9 | 0.5 | 11, 12, 20 | |
100 | 19.7 | 3.5 | 0.7 | 23, 20, 16 | ||
333 | 19.7 | 4.7 | 0.7 | 16, 18, 25 | ||
1000 | 9.7 | 1.5 | 0.3 | 8, 10, 11 | ||
2600 | 11.0 | 1.0 | 0.4 | 12 P, 10 P, 11 P | ||
5200 | 9.3 | 1.5 | 0.3 | 8 P, 9 P, 11 P | ||
NOPD | 10 | 364.7 | 42.1 | 13.0 | 321, 368, 405 | |
E. coli | DMSO | - | 59.3 | 12.4 | - | 66, 67, 45 |
test item | 33 | 54.0 | 5.0 | 0.9 | 54, 49, 59 | |
100 | 53.7 | 9.3 | 0.9 | 64, 46, 51 | ||
333 | 66.3 | 5.5 | 1.1 | 60, 70, 69 | ||
1000 | 35.3 | 3.5 | 0.6 | 32, 35, 39 | ||
2600 | 39.3 | 4.2 | 0.7 | 36 P, 38 P, 44 P | ||
5200 | 35.7 | 7.5 | 0.6 | 43 P, 36 P, 28 P | ||
4-NQO | 5.0 | 223.7 | 34.7 | 3.8 | 228, 256, 187 |
DMSO = dimethyl sulfoxide
MMNG = N-methyl-N'-nitro-N-nitrosoguanidine: 5 µg/plate in DMSO
AAC = 9-aminoacradine 100 µg/plate in DMSO
NOPD = 4-nitro-o-phenylenediamine: 10 µg/plate in DMSO
4-NQO = 4-nitroquinoline-N-oxide 5 μg/plate in DMSO
SD = standard deviation
Table 4: Results of the Prival preincubation test with metabolic activation.
Strain | Test group | Dose (µg/plate) | Mean revertants per plate | SD | Factor | Individual revertant colony counts |
TA 1535 | DMSO | - | 13.7 | 2.5 | - | 16, 14, 11 |
test item | 33 | 12.0 | 2.6 | 0.9 | 15, 11, 10 | |
100 | 10.3 | 2.1 | 0.8 | 8, 12, 11 | ||
333 | 13.0 | 1.0 | 1.0 | 13, 12, 14 | ||
1000 | 10.0 | 3.6 | 0.7 | 7, 9, 14 | ||
2600 | 16.3 | 3.2 | 1.2 | 14 P, 15 P, 20 P | ||
5200 | 10.3 | 3.5 | 0.8 | 14 B P, 10 B P, 7 B P | ||
2-AA | 2.5 | 166.0 | 47.7 | 12.1 | 111, 191, 196 | |
TA 100 | DMSO | - | 31.7 | 4.0 | - | 31, 36, 28 |
test item | 33 | 27.7 | 5.5 | 0.9 | 34, 24, 25 | |
100 | 31.0 | 4.0 | 1.0 | 31, 35, 27 | ||
333 | 26.7 | 3.2 | 0.8 | 29, 28, 23 | ||
1000 | 27.3 | 5.8 | 0.9 | 24, 34, 24 | ||
2600 | 28.7 | 3.1 | 0.9 | 32 P, 28 P, 26 P | ||
5200 | 19.3 | 1.5 | 0.6 | 18 P B, 21 P B, 19 P B | ||
2-AA | 2.5 | 639.7 | 82.9 | 20.2 | 735, 599, 585 | |
TA 1537 | DMSO | - | 6.3 | 1.5 | - | 6, 5, 8 |
test item | 33 | 7.0 | 3.0 | 1.1 | 10, 7, 4 | |
100 | 9.0 | 4.4 | 1.4 | 7, 6, 14 | ||
333 | 8.0 | 1.7 | 1.3 | 7, 10, 7 | ||
1000 | 8.7 | 2.1 | 1.4 | 8, 11, 7 | ||
2600 | 1.7 | 0.6 | 0.3 | 2 P, 1 P, 2 P | ||
5200 | 2.7 | 1.5 | 0.4 | 1 P, 3 P, 4 P | ||
2-AA | 2.5 | 134.3 | 38 | 21.2 | 172, 135, 96 | |
TA 98 | DMSO | - | 25.7 | 8.7 | - | 16, 33, 28 |
test item | 33 | 24.3 | 3.1 | 0.9 | 21, 27, 25 | |
100 | 30.3 | 4 | 1.2 | 31, 26, 34 | ||
333 | 32.3 | 7.4 | 1.3 | 38, 35, 24 | ||
1000 | 19.7 | 3.1 | 0.8 | 19, 23, 17 | ||
2600 | 11.7 | 2.5 | 0.5 | 12 P, 9 P, 14 P | ||
5200 | 5.7 | 0.6 | 0.2 | 6 P, 6 P, 5 P | ||
2-AA | 2.5 | 331.7 | 18.1 | 12.9 | 351, 315, 329 | |
CoR | 210 | 594.7 | 201.0 | 23.2 | 780, 381, 623 | |
E. coli | DMSO | - | 69.0 | 11.4 | - | 77, 56, 74 |
test item | 33 | 70.0 | 12.5 | 1.0 | 60, 66, 84 | |
100 | 73.0 | 8.5 | 1.1 | 72, 82, 65 | ||
333 | 87.0 | 12.1 | 1.3 | 85, 100, 76 | ||
1000 | 59.3 | 6.4 | 0.9 | 62, 52, 64 | ||
2600 | 67.3 | 1.5 | 1.0 | 66 P, 69 P, 67 P | ||
5200 | 55.0 | 5.3 | 0.8 | 53 P, 61 P, 51 P | ||
2-AA | 60 | 557.7 | 114.5 | 8.1 | 685, 525, 463 |
DMSO = dimethyl sulfoxide
SD = standard deviation
2-AA = 2-aminoanthracene
CoR = Congo Red 210 µg/plate in DMSO
Applicant's summary and conclusion
- Conclusions:
- The test item was not found to be mutagenic.
- Executive summary:
The potential of the test item to induce point mutations by base pair substitutions or frameshifts was evaluated in an experimental study according to the OECD Guideline 471 (1997) and the EU Method B.13 (2998): a reverse mutation (Ames) assay utilising the plate incorporation method performed on Salmonella typhimurium strains TA 98, TA 100, TA 1535, TA 1537, as well as Escherichia coli strain WP2 uvrA. The test item (33, 100, 333, 1000, 2600 and 5200 μg/plate) was evaluated for its ability to induce frameshift mutations (S. typhimurium strains TA 98 and TA 1537) and base pair substitution mutations (TA 100 and TA 1535), both with and without the presence of rat liver post-mitochondrial fraction (S9 metabolic activation). Negative (solvent and sterile) and positive controls were tested in parallel; positive controls included
2-aminoanthracene with rat liver S9 mix (2.5 µg/plate in DMSO: TA1535, TA100, TA1537, TA98; and 60 µg/plate in DMSO: E. coli WP2 uvrA), 2-aminoanthracene with hamser liver S9 mix (10 µg/plate in DMSO: TA1535, TA100, TA1537, TA98, E. coli WP2 uvrA), and without S9 mix: 9-aminoacridine (100 µg/plate in DMSO: TA1537), 4-nitro-o-phenylenediamine (10 µg/plate in DMSO: TA98), N-methyl-N'-nitro-N-nitrosoguanidine (5 µg/plate in DMSO: TA1535, TA100) and Congo Red (210 µg/plate in DMSO: TA 98). As no positive result was found in the first assay, an additional Ames assay was performed using the Pival pre-incubation method, which is specific for azo dyes, using the same test item concentrations and controls.
Precipitation of the test item was found from about 2600 μg/plate onward, both with and without S9 mix. Toxicity in bacteria was occasionally observed depending on the strain and test conditions from about 1000 μg/plate and above. The results of the negative as well as the positive controls fulfilled the acceptance criteria of this study. A relevant increase in the number of his+ or trp+ revertants was not observed in the standard plate test or in the prival preincubation test, either with or without S9 mix.Thus, under the experimental conditions of this study, the test item is considered not mutagenic in theSalmonella typhimurium/Escherichia colireverse mutation assay in the absence and the presence of metabolic activation.
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