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EC number: 246-107-8 | CAS number: 24245-27-0
- 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 cytogenicity / chromosome aberration study in mammalian cells
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 2010-12-08 to 2011-04-13
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 011
- Report date:
- 2011
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
- Version / remarks:
- 1997
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)
- Version / remarks:
- 2008
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- in vitro mammalian chromosome aberration test
Test material
- Reference substance name:
- N,N'-diphenylguanidine monohydrochloride
- EC Number:
- 246-107-8
- EC Name:
- N,N'-diphenylguanidine monohydrochloride
- Cas Number:
- 24245-27-0
- Molecular formula:
- C13H13N3.ClH
- IUPAC Name:
- N,N'-diphenylguanidine hydrochloride
Constituent 1
Method
Species / strain
- Species / strain / cell type:
- Chinese hamster lung fibroblasts (V79)
- Details on mammalian cell type (if applicable):
- CELLS USED
- Source of cells: Labor für Mutagenitätsprüfungen (LMP), Technical University Darmstadt, 64287 Darmstadt, Germany
- proliferation rate: 14 hours
- Modal number of chromosomes: 22 +/-1
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
- Periodically 'cleansed' against high spontaneous background: yes
- Metabolic activation:
- with and without
- Metabolic activation system:
- Phenobarbital/ß-naphthoflavone induce rat liver S9
- Test concentrations with justification for top dose:
- With and without S9: 9.7; 19.4; 38.8; 77.5; 155.0; 310.0; 620.0; 1240.0; 2480.0 µg/mL
Concentrations were selected based on prelim. dose-range finding assay. At the highest concentration of 2480 µg/mL precipitation was observed at the end of treatment. - Vehicle / solvent:
- - Vehicle/solvent used: DMSO
- Justification for choice of solvent/vehicle: The solvent was chosen due to its solubility properties and its relative non-toxicity to the cell cultures.
Controls
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- cyclophosphamide
- ethylmethanesulphonate
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in suspension
- Cell density at seeding: 1 x 10^4 - 6 x 10^4 cells
DURATION
- Exposure duration: 4 h
- Expression time (cells in growth medium): 14 h
- Fixation time (start of exposure up to fixation or harvest of cells): 15.5 h after start of treatment
SPINDLE INHIBITOR: Colcemid
STAIN: Giemsa
NUMBER OF REPLICATIONS: 3
METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED: Cells were fixed with a mixture of methanol and glacial acetic acid (3:1 parts, respectively). After preparation the cells were stained with Giemsa and labelled with a computer-generated random code to prevent scorer bias.
NUMBER OF METAPHASE SPREADS ANALYSED PER DOSE: at least 100 well spread metaphases per culture
Only metaphases with characteristic chromosome numbers of 22 ± 1 were included in the analysis. To describe a cytotoxic effect the mitotic index (% cells in mitosis) was determined. In addition, the number of polyploid cells in 500 metaphases per culture was determined (% polyploid metaphases; in the case of this aneuploid cell line polyploid means a near tetraploid karyotype).
DETERMINATION OF CYTOTOXICITY
- Method: reduced cell number
OTHER EXAMINATIONS:
- Determination of polyploidy: yes - Rationale for test conditions:
- According to respective guidelines
- Evaluation criteria:
- A test item is classified as non-clastogenic if:
- the number of induced structural chromosome aberrations in all evaluated dose groups is in the range of the laboratory's historical control data.
and
- no significant increase of the number of structural chromosome aberrations is observed.
A test item is classified as clastogenic if:
- the number of induced structural chromosome aberrations is not in the range of the laboratory's historical control data.
and
- either a concentration-related or a significant increase of the number of structural chromosome aberrations is observed.
Although the inclusion of the structural chromosome aberrations is the purpose of this study, it is important to include the polyploids and endoreduplications. The following criterion is valid:
A test item can be classified as aneugenic if:
- the number of induced numerical aberrations is not in the range of the laboratory's historical control data. - Statistics:
- Statistical significance was confirmed by means of the Fisher's exact test (7) (p < 0.05). However, both biological and statistical significance should be considered together. If the criteria mentioned above for the test item are not clearly met, the classification with regard to the historical data and the biological relevance is discussed and/or a confirmatory experiment is performed.
Results and discussion
Test results
- Key result
- Species / strain:
- Chinese hamster lung fibroblasts (V79)
- Metabolic activation:
- with and without
- Genotoxicity:
- positive
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- at 155 µg/mL and above
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not examined
- Positive controls validity:
- valid
- Additional information on results:
- TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no
- Effects of osmolality: no
- Evaporation from medium: no
- Water solubility: the substance is considered to be well soluble in water
- Precipitation: yes, at concentrations of 2480 µg/mL and above, with and without S9
RANGE-FINDING/SCREENING STUDIES: yes
Any other information on results incl. tables
The test item, dissolved in DMSO, was assessed for its potential to induce structural chromosome aberrations in V79 cells of the Chinese hamster in vitro in the absence and presence of metabolic activation by S9 mix. According to the OECD Guideline only one experiment was performed, since the test item was considered to be clastogenic after 4 hours treatment. The chromosomes were prepared 18 hours after start of treatment with the test item. The exposure period was 4 hours with and without metabolic activation. In each experimental group two parallel cultures were set up. 100 metaphases per culture were evaluated for structural chromosome aberrations. No relevant influence of the test item on pH value or osmolarity was observed (solvent control 398 mOsm, pH 7.43 versus 389 mOsm and pH 7.45 at 2480.0 µg/mL). Precipitation of the test item in culture medium was observed after 4 hours treatment with 2480.0 µg/mL and above in the presence and absence of S9 mix. Concentrations between 9.7 and 2480.0 ug/mL were applied. Clear cytotoxic effects were observed after 4 hours treatment with 155.0 µg/mL and above in the absence of S9 mix, where the cell numbers were reduced to about 60 % of solvent control cells. In the presence of S9 mix dose groups showing relevant cytotoxic effects were not evaluable for cytogenetic damage. In the absence of S9 mix a dose-dependent increase in the number of aberrant cells, excluding gaps was observed (5.5, 7.0 and 16.0 %, respectively) after treatment with 38.8, 77.5 and 155.0 µg/mL. All values exceeded the total range of the laboratory's historical solvent control (0.0 - 4.0 % aberrant cells, excluding gaps). In addition, the value obtained at 155.0 µg/mL (16.0 % aberrant cells, excluding gaps) was statistically significantly increased. In the presence of S9 mix a statistically significant and dose-dependent increase in the number of aberrant cells (5.5 %) was observed at the highest evaluable concentration (310.0 µg/mL). This statistically significant value also exceeded the total range of the laboratory's historical solvent control (0.0 - 4.0 % aberrant cells, excluding gaps). No relevant evidence of an increase in polyploid metaphases was noticed after treatment with the test item as compared to the controls. Therefore, biologically relevant increases of chromosomal aberrations were observed in the absence and presence of metabolic activation and the test item is thus considered clastogenic under the conditions of this study.
Table 1: Summary of results of the chromosome aberration study with the test item
* Inclusive cells carrying exchanges
Preparation interval |
Test item concentration in µg/mL |
Polyploid cells in % |
Cell numbers in% of control |
Mitotic indices in % of control |
Incl. gaps* |
Aberrant cells in % excl. gaps* |
with exchanges |
||||||
Exposure period 4 hrs without S9 mix |
|||||||||||||
18 hrs |
Solvent control1 |
2.7 |
100.0 |
100.0 |
4.5 |
3.5 |
1.0 |
||||||
Positive control2 |
1.7 |
n.t. |
88.1 |
44.0 |
44.0S |
33.5 |
|||||||
38.8 |
2.1 |
92.0 |
106.0 |
6.0 |
5.5 |
3.0 |
|||||||
77.5 |
2.4 |
95.5 |
111.5 |
7.0 |
7.0 |
3.5 |
|||||||
155.0 |
2.5 |
60.4 |
84.5 |
16.5 |
16.0S |
5.0 |
|||||||
Exposure period 4 hrs with S9 mix |
|||||||||||||
18 hrs |
Solvent control1 |
2.0 |
100.0 |
100.0 |
3.0 |
2.0 |
0.5 |
||||||
Positive control3 |
2.1 |
n.t. |
67.6 |
21.5 |
20.5S |
6.5 |
|||||||
77.5 |
1.8 |
106.5 |
107.3 |
2.5 |
2.5 |
1.5 |
|||||||
155.0 |
3.0 |
108.5 |
84.6 |
5.0 |
3.5 |
1.0 |
|||||||
310.0 |
2.9 |
100.0 |
95.8 |
6.0 |
5.5 |
1.5 |
|||||||
n.t. Not tested
S Aberration frequency statistically significant higher than corresponding control values
1 DMSO 0.5 % (v/v)
2 EMS 1000.0 µg/mL
3 CPA 1.4 µg/mL
Applicant's summary and conclusion
- Conclusions:
- The test item is considered to be clastogenic in this chromosome aberration test in the absence and presence of metabolic activation.
- Executive summary:
The test item, dissolved in DMSO, was assessed for its potential to induce structural chromosome aberrations in V79 cells of the Chinese hamster in vitro in one experiment. The following study design was performed:
Without S9 mix
With S9 mix
Exposure period
4 h
4 h
Recovery
14 h
14 h
Preparation interval
18 h
18 h
In each experimental group two parallel cultures were set up. At least 100 metaphases per culture were evaluated for structural chromosome aberrations. The highest applied concentration (2480.0 µg/mL; approx. 10.0 mM) was chosen with regard to the molecular weight of the test item and with respect to the current OECD Guideline 473. Dose selection for the cytogenetic experiments was performed considering the toxicity data. In the absence and presence of S9 mix concentrations higher than the evaluated were not evaluable for cytogenetic damage due to exceedingly strong cytotoxic effects. The cell numbers were reduced to approximately 60 % in the highest evaluable dose group in the experimental part without S9 mix. In the absence of S9 mix a dose-dependent increase in the number of aberrant cells, excluding gaps was observed (5.5, 7.0 and 16.0 %%, respectively) after treatment with 38.8, 77.5 and 155.0 µg/mL All values exceeded the total range of the laboratory's historical solvent control (0.0 - 4.0 % aberrant cells, excluding gaps). In addition, the value obtained at 155.0 µg/mL (16.0 % aberrant cells, excluding gaps) was statistically significantly increased. In the presence of S9 mix a statistically significant and dose-dependent increase in the number of aberrant cells (5.5 %) was observed at the highest evaluable concentration (310.0 µg/mL). This value exceeded the total range of the laboratory's historical solvent control (0.0 - 4.0 % aberrant cells, excluding gaps) and is also statistically significant. No relevant evidence of an increase in polyploid metaphases was found after treatment with the test item as compared to the control cultures. Appropriate mutagens were used as positive controls. They induced statistically significant increases (p < 0.05) in cells with structural chromosome aberrations.
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