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EC number: 934-405-7 | 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:
- 2016
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 016
- Report date:
- 2016
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Version / remarks:
- 9th addendum
- 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
Test material
- Test material form:
- solid: particulate/powder
Constituent 1
- Specific details on test material used for the study:
- SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material:040802#
- Expiration date of the lot/batch:06.08.2016
- Purity test date:08/01/2015
- Purity: 99.17%
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material:room temperature
- Stability under test conditions:stable
- Solubility and stability of the test substance in the solvent/vehicle:stable
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium:none
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing:the test item is dissolved in Aqua destillata and diluted prior to treatment. Aqua distilla is compatible with bacteria and the S9 activity.
- Final dilution of a dissolved solid, stock liquid or gel:different concentration tested (see below)
FORM AS APPLIED IN THE TEST (if different from that of starting material): liquid
Method
Species / strain
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
- Details on mammalian cell type (if applicable):
- TA 98:
his D 3052; rfa-; uvrB-; R-factor: frame shift mutations
TA 100:
his G 46; rfa-; uvrB-; R-factor: base-pair substitutions
TA 1535:
his G 46; rfa-; uvrB-: base-pair substitutions
TA 1537:
his C 3076; rfa-; uvrB-: frame shift mutations
TA 102:
his G 428 (pAQ1); rfa-; R-factor: base-pair substitutions
Tester strains TA 98, TA 1535 and TA 102 were obtained from MOLTOX, INC., NC 28607, USA.
Tester strains TA 100 and TA 1537 were obtained from Xenometrix AG, Switzerland. They were
stored as stock cultures in ampoules with nutrient broth (OXOID) supplemented with DMSO (approx.
8% v/v) over liquid nitrogen.
All Salmonella strains contain mutations in the histidine operon, thereby imposing a requirement for
histidine in the growth medium. They contain the deep rough (rfa) mutation, which deletes the
polysaccharide side chain of the lipopolysaccharides of the bacterial cell surface. This increases cell
permeability of larger substances. The other mutation is a deletion of the uvrB gene coding for a
protein of the DNA nucleotide excision repair system resulting in an increased sensitivity in detecting
many mutagens. This deletion also includes the nitrate reductase (chl) and biotin (bio) genes
(bacteria require biotin for growth).
The tester strains TA 98, TA 100 and TA 102 contain the R-factor plasmid, pkM101. These strains
are reverted by a number of mutagens that are detected weakly or not at all with the non R-factor
parent strains. pkM101 increases chemical and spontaneous mutagenesis by enhancing an errorprone
DNA repair system which is normally present in these organisms.
The properties of the S. typhimurium strains with regard to membrane permeability, ampicillin- and
tetracycline-resistance as well as normal spontaneous mutation rates are checked regularly
according to Ames et al.In this way it is ensured that the experimental conditions set up by
Ames are fulfilled.
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9 mix
- Test concentrations with justification for top dose:
- EXPERIMENT I (Plate incorporation method) 3.16, 10.0, 31.6, 100, 316, 1000 and 2500 µg/plate and 1.00 µg/plate only for TA 1535 and TA1537
EXPERIMENT II (pre incubation method) 1.00, 3.16, 10.0, 31.6, 100, 316, 1000 and 2500 µg/plate
Justification of top dose: for soluble non toxic test compounds the recommended maximum test concentration is 5 µl/plate and a pre-experiment test was performed for toxicity evaluation showing no background lawn from 2500 µg/plate in one strain - Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: purified water (Aqua Destillata)
- Justification for choice of solvent/vehicle:The solvent was compatible with the survival of the bacteria and the S9 activity.
Controls
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Remarks:
- The solvent and the negative controls were the same.
- True negative controls:
- no
- Positive controls:
- yes
- Remarks:
- 4 different positive controls depending on the metabolic activation or not and the tester strains.
- Positive control substance:
- sodium azide
- methylmethanesulfonate
- other: 4-nitro-o-phenylene-diamine and 2-aminoanthracene
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in medium
- Cell density at seeding (if applicable):approx. 10^9 cells/ml
DURATION
- Preincubation period:60 min in experiment II
- Exposure duration:48h for both experiments
NUMBER OF REPLICATIONS:3
DETERMINATION OF CYTOTOXICITY
- Method: detected by a clearing / diminution of the background lawn or a reduction in the number of revertants down to a mutation factor of 0.5 or less in relation to the solvent control
OTHER EXAMINATIONS:
colonies counted using a ProtoCOL counter (Meintrup DWS Laborgäte GmbH) or manually (for TA 1535 and 1537 or if precipitation occurred)
- OTHER:- preparation of bacteria: each strain was grown by culturing for 12h at 37°C in Nutrient Broth (8g/l Nutrient Broth + 5 g/l NaCl). Ampicillin (10 mg/ml) was added to TA 98, TA 100 and TA 102.
- Agar plates: Vogel-Bonner Medium E agar plates with 2% glucose, sterilized 20 min at 121°C in autoclave.
- Overlay agar: contains agar-agar, NaCl, LhistidinexHClxH2O, biotin, sterilized 20 min at 121°C in autoclave
- Metabolic activation system: S9 liver microsomal fraction (from Male Wistar rats and Sprague Dawley rats (phenobarbital/Bêta-naphthoflavone)) in a S9 mix preparation with MgCl2, KCl, glucose-6-phosphate and NADP, stored on ice.
- For the test without metabolic activation, S9 mix is replaced by a sterilized phosphate buffer solution stored at 4°C - Evaluation criteria:
- Mutation factor is calculated by dividing the mean value of the revertant counts by the mean values of the solvent control.
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 at least one tester strain with or without metabolic activation.
A biologically relevant increase is described as follows:
- if in tester strains TA 98, TA 100 and TA 102 the number of reversions is at least twice as high
- if in tester strains TA 1535 and TA 1537 the number of reversions is at least three times higher than the reversion rate of the solvent control - Statistics:
- Not necessary because the biological relevance of the results is the criterion for the interpretation of the results
Results and discussion
Test resultsopen allclose all
- Key result
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- concentration with toxic effects depends on experimental conditions
- Vehicle controls validity:
- valid
- Untreated 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:
- cytotoxicity
- Remarks:
- concentration with toxic effects depends on experimental conditions
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 102
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- concentration with toxic effects depends on experimental conditions
- Vehicle controls validity:
- valid
- Untreated 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:
- cytotoxicity
- Remarks:
- concentration with toxic effects depends on experimental conditions
- Vehicle controls validity:
- valid
- Untreated 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:
- cytotoxicity
- Remarks:
- concentration with toxic effects depends on experimental conditions
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Additional information on results:
- TEST-SPECIFIC CONFOUNDING FACTORS
-Precipitation: no precipitation observed
HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Negative (solvent/vehicle) historical control data:
mean values of the spontaneous reversion frequency are within the historical control data range(2012 -2014):
- S9 + S9
min max min max
TA 98 13 48 13 61
TA 100 61 182 68 194
TA 1535 4 35 4 34
TA 1537 2 27 3 31
TA 102 136 415 91 495
Any other information on results incl. tables
Concentrations were toxic effects where observed:
EXPERIMENT I | EXPERIMENT II | |||
with (+S9) | without (-S9) | with (+S9) | without (-S9) | |
TA98 | ≥1000 | ≥1000 | 2500 | ≥ 316 |
TA100 | ≥ 1000 | ≥ 1000 | ≥ 1000 | 2500 |
TA102 | ≥ 1000 | ≥1000 | ≥ 316 | ≥ 316 |
TA1535 | ≥ 1000 | 2500 | ≥ 1000 | ≥ 316 |
TA1537 | 2500 | ≥ 100 | ≥ 316≥ | ≥ 100 |
No biologically relevant increases in revertant colony numbers of any of the five tester strains were observed following treatment with X330 at any concentration level, neither in the presence nor absence of metabolic activation in experiment I and II.
The reference mutagens induced a distinct increase of revertant colonies indicating the validity of the experiments.
Applicant's summary and conclusion
- Conclusions:
- Under these experimental conditions and during the described mutagenicity test, X330 did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used. Therefore X330 is considered to be non mutagenic in this bacterial reverse mutation assay.
- Executive summary:
In order to investigate the potential of X330 for its ability to induce gene mutations the plate incorporation test (experiment I) and the pre-incubation test (experiment II) were performed with the Salmonella typhimurium strains TA 98, TA 100, TA 1535, TA 1537 and TA 102.
In two independent experiments several concentrations of the test item were used. Each assay was conducted with and without metabolic activation.
No precipitation of the test item was observed in any tester strain used in experiment I and II (with and without metabolic activation).
No biologically relevant increases in revertant colony numbers of any of the five tester strains were observed following treatment with X330 at any concentration level, neither in the presence nor absence of metabolic activation in experiment I and II.
The reference mutagens induced a distinct increase of revertant colonies indicating the validity of the experiments.
In conclusion, under these experimental conditions and during the described mutagenicity test, X330 did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used. Therefore X330 is considered to be non mutagenic in this bacterial reverse mutation assay.
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