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EC number: 700-812-1 | CAS number: 1189052-95-6
- 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
Link to relevant study records
- Endpoint:
- in vitro cytogenicity / chromosome aberration study in mammalian cells
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- Arrival of the Test Item: 16 June 2011 Date of Final Report: 24 February 2012
- Reliability:
- 1 (reliable without restriction)
- Reason / purpose for cross-reference:
- reference to same study
- Reason / purpose for cross-reference:
- reference to other study
- Qualifier:
- according to guideline
- Guideline:
- other: OECD 487
- Deviations:
- yes
- Remarks:
- See report for details. deviation did not affect quality of the study
- Principles of method if other than guideline:
- Ninth Addendum to OECD Guidelines for Testing of Chemicals, Section 4, No. 487, ‘’In Vitro Mammalian Cell Micronucleus Test’’, adopted 22nd July, 2010.
Kirsch-Volders M. et al., “Report from in vitro micronucleus assay working group”, (2003), Mut. Res. 540, 153 – 163.
Kirsch-Volders M. et al., “Report from the in vitro micronucleus assay working group”, (2000), Environm. Mol. Mut. 35, 167 - 172.
Von der Rude W. et al, “In vitro micronucleus assay with Chinese hamster V79 cells results of a collaborative study with in situ exposure to 26 chemical substances”, (2000), Mut. Res, 468, 137- 163.
Fenech M., “Cytokinesis-block micronucleus cytome assay”, (2007), Nature Protocols 2 (5), 1084-1104. - GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- mammalian cell gene mutation assay
- Target gene:
- See attached report
- Species / strain / cell type:
- Chinese hamster lung fibroblasts (V79)
- Additional strain / cell type characteristics:
- not applicable
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9
- Test concentrations with justification for top dose:
- Exposure Concentrations: The selection of the test item concentrations used and evaluated in the experiments based on cytotoxicity of the test item that intensified with increasing test item concentrations and occurred at concentrations where a low and constant level of precipitation was observed.
Duplicate cultures were treated at each concentration. The following concentrations were used in the main experiments:
Experiment I:
without metabolic activation: 750, 1000, 1200, 1300, 1400, 1500, 1550, 1600, 1650 and 1700 µg/mL
with metabolic activation: 1500, 1600, 1650, 1675, 1700, 1725, 1750, 1775 and 1800 µg/mL
Experiment II:
without metabolic activation: 31.6, 100, 316, 1000, 1250, 1500, 1600, 1700, 1800, 1900, 2000, 2250 and 2500 µg/mL
The following concentrations were selected for the microscopic analyses:
Experiment I with short exposure (4 h):
without metabolic activation: 1200, 1300, 1400 and 1500 µg/mL with metabolic activation: 1500, 1600, 1650 and 1675 µg/mL
Experiment II with extended exposure (20 h):
without metabolic activation: 1250, 1500, 1600 and 1700 µg/mL - Vehicle / solvent:
- MEM
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- ethylmethanesulphonate
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- cyclophosphamide
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- other: Colecimide
- Details on test system and experimental conditions:
- Test System
Cells: V79 cells in vitro have been widely used to examine the ability of chemicals to induce cytogenetic changes and thus identify potential carcinogens or mutagens, These cells are chosen because of their stable karyotype and their low spontaneous induction rate of micronucleus formation under standardized culture conditions. These facts are necessary for the appropriate performance of the study.
The V79 cells (ATCC, CCL-93) are stored over liquid nitrogen (vapour phase) in the cell bank of BSL BIOSERVICE, as large stock cultures allowing the repeated use of the same cell culture batch in experiments. Routine checking of mycoplasma infections were carried out before freezing.
For the experiments thawed cultures are set up in 75 cm2 cell culture plastic flasks at 37°C in a 5% carbon dioxide atmosphere (95% air). 5 x 105 cells per flask are seeded in 15 mL of MEM (minimum essential medium) supplemented with 10% FBS (foetal bovine serum) and subcultures are made every 3-4 days.
Mammalian Microsomal Fraction S9 Homogenate: An advantage of using in vitro cell cultures is the accurate control of the concentration and exposure time of cells to the test item under study. However, due to the limited capacity of cells growing in vitro for metabolic activation of potential mutagens, an exogenous metabolic activation system is necessary. Many substances only develop mutagenic potential when they are metabolized by the mammalian organism. Metabolic activation of substances can be achieved by supplementing the cell cultures with liver microsome preparations (S9 mix).
The S9 liver microsomal fraction was prepared at BSL BIOSERVICE GmbH. Male Wistar rats were induced with phenobarbital (80 mg/kg bw) and B-naphthoflavone (100 mg/kg bw) [10,11] for three consecutive days by oral route.
The following quality control determinations are performed:
a) Biological activity in:
- the Salmonella typhimurium assay using 2-aminoanthracene
- the mouse lymphoma assay using benzo[a]pyrene
- the chromosome aberration assay using cyclophosphamide.
b) Sterility Test
A stock of the supernatant containing the microsomes was frozen in aliquots of 2 and 4.5 mL and stored at ≤-75°C.
The protein concentration in the S9 preparation (Lot: 210711) was 38.5 mg/mL.
S9 Mix: An appropriate quantity of the S9 supernatant was thawed and mixed with S9 cofactor solution to result in a final protein concentration of 0.75 mg/mL in the cultures. Cofactors were added to the S9 mix to reach the concentrations below:
8 mM MgCl2
33 mM KCl
5 mM Glucose-6-phosphate
5 mM NADP
in 100 mM sodium-phosphate-buffer pH 7.4. During the experiment the S9 mix was stored on ice.
Culture Medium:
Complete Culture Medium: MEM medium supplemented with:
10 % foetal bovine serum (FBS)
100 U/100 µg/mL penicillin/streptomycin solution
2 mM L-glutamine
2.5 µg/mL amphotericin
25 µM HEPES
Treatment Medium (short-time exposure): Complete culture medium without FBS.
After Treatment Medium / Treatment Medium (long-time exposure): Complete culture medium with 10% FBS and 1.5 µg/mL cytochalasin B.
(10). Elliot, EM., Combes, RD., Elcombe, CR., Gatehouse, DO., Gibson, G.G., Mackay, J.M. and Wolf, R.C. (1992). Report of UK Environmental Mutagen Society Working Party. Alternatives to Aroclor 1254- induced S9 in In Vitro Metabolic Activation in Mutagenesis Testing Mutagenesis, 7, 175-177
(11). Matsushima, T., Sawamura, M., Hara, K. and Sugimura, T. (1976). A Safe Substitute for Polychlorinated Biphenyls as an Inducer of Metabolic Activation Systems. In: de Serres, F.)., Fouts, JR., Bend, J.R. and Philpot R.M., (eds). In Vitro Metabolic Activation in Mutagenesis Testing Elsevier, North-Holland, 85-88 - Evaluation criteria:
- See Attached report
- Statistics:
- Not used in the analysis
- Species / strain:
- Chinese hamster lung fibroblasts (V79)
- 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
- Additional information on results:
- See attached report
- Remarks on result:
- other: all strains/cell types tested
- Remarks:
- Migrated from field 'Test system'.
- Conclusions:
- Interpretation of results (migrated information):
negative
In conclusion, it can be stated that during the study described and under the experimental conditions reported, FHP-OHS did not induce structural and/or numerical chromosomal damage in Chinese hamster V79 cells.
Therefore, FHP-OHS is considered to be non-mutagenic with respect to clastogenicity and/or aneugenicity in this in vitro Mammalian Cell Micronucleus Test. - Executive summary:
Summary Results: In order to investigate FHP-OHS for a possible potential to induce micronuclei in Chinese hamster V79 cells an in vitro a micronucleus assay was carried out.
The test item was dissolved in cell culture medium, treated with ultrasound for 15 minutes at 37°C and diluted within 1 hour prior to treatment. The pH-value detected with the test item was within the physiological range (7.0-7.4).
The selection of the concentrations used in experiment I and II was based on data from the pre-experiment according to the guidelines.
In experiment I 1500µg/mL without metabolic activation and 1675µg/mL with metabolic activation was selected as highest dose group for the microscopic analysis of micronuclei.
In experiment II without metabolic activation 1700µg/mL was selected as highest dose group for the microscopic analysis of micronuclei.
The following concentrations were evaluated:
Experiment I with short exposure (4 h):
without metabolic activation: 1200, 1300, 1400 and 1500µg/mL with metabolic activation: 1500, 1600, 1650 and 1675µg/mL
Experiment II with extended exposure (20 h):
without metabolic activation: 1250,1500, 1600 and 1700µg/mL
The test item was prepared in culture medium (MEM medium). A low level of precipitation was observed at all concentrations used and evaluated in experiment I and II that did not intensify with increasing test item concentrations.
In experiment I without metabolic activation no decrease of the relative CBPI below 70 % was noted up to a concentration of 1400µg /ml. At a concentration of 1500µg /mL a relative CBPI of 57% was observed. In experiment I with metabolic activation no decrease of the relative CBPI below 70 % was noted up to a concentration of 1600µg/ml. At a concentration of 1650µg/mL a relative CBPI of 66% and at a concentration of 1675µg/mL a relative CBPI of 52% was noted. In experiment II without metabolic activation no decrease of the relative CBPI below 70% was noted up to a concentration of 1500µg/mL. At a concentration of 1600µg/mL a relative CBPI of 67% and at a concentration of 1700µg/mL a relative CBPI of 48% was noted.
The micronucleus frequencies found in the groups treated with the test item did not show a biologically relevant increase compared to the corresponding negative control.
The nonparametric x2 Test was performed to verify the results in both experiments. No statistically significant enhancement (p<0.05) of cells with micronuclei was noted in the dose groups of the test item evaluated in experiment I and II without metabolic activation (Table 9-11 of attached report). In experiment I with metabolic activation the micronucleus frequencies at concentrations of 1600µg/mL and 1675µg/mL were statistically significantly increased compared to the corresponding negative control (as determined by the nonparametric x2 Test), However, these micronucleus frequencies were within the range of the historical negative control data and no dose-response relationship could be observed. Based on this data the observed increases were regarded as not biologically relevant.
Ethylmethanesulfonate (, 900 and 600µg/mL) and Cyclophosphamide (CPA, 2.5µg/mL) were used as clastogenic controls. Colcemide (0.08 and 0.8µg/mL) was used as aneugenic control. All induced distinct and biologically relevant increases of micronucleus frequency. This demonstrates the validity of the assay.
Reference
Pre-Experiment for Toxicity: According to OECD guideline 487 the highest recommended dose is 5000µg/mL. The test item was dissolved in MEM medium and treated with ultrasound for 15 minutes at 37°C. Precipitation of the test item was noted at a low and constant level at concentrations of 500µg/mL and higher (without metabolic activation) and at concentrations of 31.3µg/mL and higher (with metabolic activation) at the end of treatment. The highest dose group evaluated in the pre-experiment was 1000µg/mL. The cytokinesis block proliferation index (CBPI) was used as parameter for toxicity. The concentrations evaluated in the main experiment based on the results obtained in the pre-experiment (Table 1 of attached report).
Precipitation: The test item was prepared in culture medium (MEM medium). A low level of precipitation was observed at all concentrations used and evaluated in experiment I and II that did not intensify with increasing test item concentrations.
Toxicity: In experiment I without metabolic activation no decrease of the relative CBPI below 70 % was noted up to a concentration of 1400µg/ml. At a concentration of 1500µg/mL a relative CBPI of 57% was observed. In experiment I with metabolic activation no decrease of the relative CBPI below 70 % was noted up to a concentration of 1600µg/ml. At a concentration of 1650µg/mL a relative CBPI of 66% and at a concentration of 1675µg/mL a relative CBPI of 52% was noted.
In experiment II without metabolic activation no decrease of the relative CBPI below 70% was noted up to a concentration of 1500µg/mL. At a concentration of 1600µg/mL a relative CBPI of 67% and at a concentration of 1700µg/mL a relative CBPI of 48% was noted. |
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Additional information
Justification for selection of genetic toxicity endpoint
Most recent study
Justification for classification or non-classification
Negative results form both studies
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