Registration Dossier
Registration Dossier
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EC number: 202-993-8 | CAS number: 101-97-3
Endpoint summary
Administrative data
Key value for chemical safety assessment
Genetic toxicity in vitro
Description of key information
An Ames test and a test on mammalian cell lines was conducted to determine the mutagenic potential of the test item. The test item showed no mutagenic potential in both tests.
The test item was assessed for its potential
to induce structural chromosomal aberrations in human lymphocytes in
vitro according to OECD guideline 473. The test item did not induce
structural chromosomal aberrations.
The HPRT test was performed to investigate the potential of the test
item to induce gene mutations at the HPRT locus in V79 cells. It can be
stated that under the experimental conditions reported the test item did
not induce gene mutations at the HPRT locus in V79 cells.
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:
- 2015-01-14 to 2015-07-06
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
- Version / remarks:
- 2014
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 870.5375 - In vitro Mammalian Chromosome Aberration Test
- Version / remarks:
- 1998
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- in vitro mammalian chromosome aberration test
- Species / strain / cell type:
- lymphocytes: human
- Metabolic activation:
- with and without
- Metabolic activation system:
- Phenobarbital/β-naphthoflavone induced rat liver S9 was used as the metabolic activation system
- Test concentrations with justification for top dose:
- 10.67, 18.67, 32.67, 57.17, 100.04, 175.07, 306.38, 536.16, 938.29, 1642.0 µg/mL (Exp I and IIA without S9 Mix)
100.04, 175.07, 201.11, 268.15, 306.38, 402.22, 536.29, 670.36 µg/mL (Exp. IIB without S9 Mix)
10.67, 18.67, 32.67, 57.17, 100.04, 175.07, 306.38, 536.16, 938.29, 1642.0 µg/mL (Exp I with S9 Mix)
100.04, 175.07, 306.38, 536.16, 938.29, 1642.0 µg/mL (Exp IIA with S9 Mix) - Vehicle / solvent:
- DMSO
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- ethylmethanesulphonate
- Remarks:
- Without metabolic activation
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- cyclophosphamide
- Remarks:
- With metabolic activation
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in medium
DURATION
- Exposure duration: Without S9-Mix: 4 h, 22 h
With S9 Mix: 4 h
- Fixation time: The cultures were harvested by centrifugation 22 h after beginning of treatment
SPINDLE INHIBITOR: Colcemid (final concentration: 0.2 μg/mL)
STAIN: Giemsa
NUMBER OF REPLICATIONS: 2
NUMBER OF CELLS EVALUATED: At least 150 metaphases per culture were evaluated
DETERMINATION OF CYTOTOXICITY
- Method: Percentages of mitotic suppression in comparison with the controls by counting 1000 cells per culture in duplicate - Evaluation criteria:
- The chromosomal aberration assay will be considered acceptable if it meets the following criteria:
a) The rate of chromosomal aberrations in the solvent controls falls within the historical laboratory control data range.
b) The rate of chromosomal aberrations in the positive controls is statistically significant increased.
A test item can be classified as non-clastogenic if:
− the number of induced structural chromosomal aberrations in all evaluated dose groups is in the range of the historical laboratory control data and
− no statistically significant increase of the rate of structural chromosomal aberrations is observed in comparison to the respective solvent control.
A test item can be classified as clastogenic if:
− the number of induced structural chromosomal aberrations is not in the range of the historical laboratory control data and
− either a concentration-related or a statistically significant increase in the number of cells carrying structural chromosomal aberrations is observed. - Statistics:
- The current historical data range together with the statistical significance, confirmed by the Fisher’s exact test (modified) (p < 0.05) using the validated R Script LM.Rnw, should be considered for classification of the test item.
- Key result
- Species / strain:
- lymphocytes: human
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not examined
- Positive controls validity:
- valid
- Additional information on results:
- The test item, dissolved in DMSO, was assessed for its potential to induce chromosomal aberrations in human lymphocytes in vitro in the absence and presence of metabolic activation by S9 mix.
Three independent experiments were performed. In Experiment I the exposure period was 4 hours with and without S9 mix. In Experiment IIA the exposure period was 4 hours with S9 mix and 22 hours without S9 mix. In Experiment IIB (confirmatory experiment) the exposure period was 22 hours without S9 mix. The chromosomes were prepared 22 hours (in all experiments) after start of treatment with the test item.
In each experimental group two parallel cultures were analysed. At least 150 metaphases per culture were evaluated for structural chromosomal aberrations, except for the positive control in Experiment II, in the absence of S9 mix, where only 50 metaphases were evaluated due to strong clastogenic effects. 1000 cells were counted per culture for determination of the mitotic index.
The highest treatment concentration in this study, 1642.0 µg/mL (approx. 10 mM) was chosen with regard to the molecular weight of the test item and with respect to the OECD Guideline for in vitro mammalian cytogenetic tests.
In Experiment I and IIA, phase separation of the test item in the culture medium was observed at 536.16 µg/mL and above in the absence and presence of S9 mix at the end of treatment.
No relevant influence on osmolarity or pH was observed.
In Experiment I in the absence and presence of S9 mix, in Experiment IIB in the absence of S9 mix and in Experiment IIA in the presence of S9 mix, no cytotoxicity was observed up to the highest applied concentration. In Experiment IIA in the absence of S9 mix no cytotoxicity was observed up to the highest evaluated concentration. Toxic effects, indicated by reduced mitotic indices were observed at higher concentrations.
In Experiments I and IIB, in the absence of S9 mix and Experiments I and IIA in the presence of S9 mix, no statistically significant or biologically relevant increase in the number of cells carrying structural chromosome aberrations was observed. The aberration rates of the cells after treatment with the test item (0.3 – 3.0% aberrant cells, excluding gaps) exceeded the range of the solvent control values (1.0 – 2.3% aberrant cells, excluding gaps), but were within the range of the laboratory historical solvent control data. In Experiment IIA in the absence of S9 mix a minor but statistically significant increase was observed, following continuous treatment. Therefore a confirmatory experiment with closer concentrations was performed (reported as Exp. IIB). Since no increase was observed in this confirmatory experiment, the observed statistical significant values are judged as biological irrelevant.
Either EMS (660.0 or 770.0 µg/mL) or CPA (7.5 or 15.0 µg/mL) were used as positive controls and showed distinct increases in cells with structural chromosome aberrations. - 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 under the experimental conditions reported, the test item did not induce structural chromosomal aberrations in human lymphocytes in vitro.
Therefore, the test item is considered to be non-clastogenic in this chromosome aberration test, when tested up to insoluble concentrations (phase separation) and in Experiment IIA up to the highest evaluable concentration. - Executive summary:
The test item, dissolved in DMSO, was assessed for its potential to induce structural chromosomal aberrations in human lymphocytes in vitro in three independent experiments. 150 metaphases were evaluated for structural chromosomal aberrations, except for the positive control in Experiment II, in the absence of S9 mix, where only 50 metaphases were evaluated.
The highest applied concentration in this study (1642.0 µg/mL of the test item, approx. 10 mM) was chosen with regard to the molecular weight of the test item and with respect to the current OECD Guideline 473.
Dose selection of the cytogenetic experiment was performed considering the toxicity data and the solubility of the test item in accordance with the OECD Guideline 473. In Experiment I in the absence and presence of S9 mix, in Experiment IIB in the absence of S9 mix and in Experiment IIA in the presence of S9 mix, no cytotoxicity was observed up to the highest applied concentration. In Experiment IIA in the absence of S9 mix no cytotoxicity was observed up to the highest evaluated concentration. Toxic effects, indicated by reduced mitotic indices were observed at higher concentrations.
In Experiments I and IIB, neither a statistically significant nor a biologically relevant increase in the number of cells carrying structural chromosomal aberrations was observed after treatment with the test item. Only in Experiment IIA a minor but statistically significant increase was observed in the absence of S9 mix following continuous treatment, therefore a confirmatory experiment with closer concentrations was performed (reported as Exp. IIB).
No evidence of an increase in polyploid metaphases was noticed after treatment with the test item as compared to the control cultures. Appropriate mutagens were used as positive controls. They induced statistically significant increases in cells with structural chromosome aberrations. In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce structural chromosomal aberrations in human lymphocytes in vitro.
Therefore, is considered to be non-clastogenic in this chromosome aberration test, when tested up to insoluble concentrations (phase separation) and in Experiment IIA up to the highest evaluable concentration.
- Endpoint:
- in vitro gene mutation study in mammalian cells
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2015-11-10 to 2016-01-26
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
- Version / remarks:
- 2015
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)
- Version / remarks:
- 2008
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 870.5300 - In vitro Mammalian Cell Gene Mutation Test
- Version / remarks:
- 1998
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- mammalian cell gene mutation assay
- Species / strain / cell type:
- Chinese hamster lung fibroblasts (V79)
- Metabolic activation:
- with and without
- Metabolic activation system:
- Phenobarbital/β-naphthoflavone induced rat liver S9
- Test concentrations with justification for top dose:
- 25.6, 51.3, 102.5, 205.0, 410.0, 820.0 µg/mL
- Vehicle / solvent:
- DMSO
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- ethylmethanesulphonate
- Remarks:
- Without metabolic activation
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- 7,12-dimethylbenzanthracene
- Remarks:
- With metabolic activation
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in medium
DURATION
- Exposure duration: 4 hours
- Expression time: Approx. 7 days
- Fixation time: Approx. 8 days
SELECTION AGENT: 11 μg/mL 6-thioguanine.
DETERMINATION OF CYTOTOXICITY
- Method: cloning efficiency - Evaluation criteria:
- A test item is classified as positive if it induces a concentration-related increase of the mutant frequency exceeding the historical solvent control range.
A test item producing no concentration-related increase of the mutant frequency above the historical solvent control range is considered to be non-mutagenic in this system.
A mutagenic response is described as follows:
The test item is classified as mutagenic if it induces with at least one of the concentrations in both parallel cultures a mutation frequency that exceeds the historical negative and solvent control data range (95% confidence interval limits).
The increase should be significant and dose dependent as indicated by statistical analysis (linear regression, least squares).
However, in a case by case evaluation this decision depends on the level of the corresponding solvent control data. If there is by chance a low spontaneous mutation rate within the laboratory´s historical control data range, a concentration-related increase of the mutations within this range has to be discussed. The variability of the mutation rates of solvent controls was also taken into consideration. - Statistics:
- A linear regression was performed to assess a possible dose dependent increase of mutant frequencies. The numbers of mutant colonies generated with the test item were compared to the solvent control groups. A trend is judged as significant whenever the p-value (probability value) is below 0.05. However, both, biological and statistical significance was considered together.
- Key result
- 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:
- not examined
- Positive controls validity:
- valid
- Additional information on results:
- No relevant cytotoxic effect indicated by an adjusted relative cloning efficiency below 50% in both cultures occurred up to the maximum concentration with and without metabolic activation.
No relevant and reproducible increase in mutant colony numbers/106 cells was observed in the main experiment up to the maximum concentration. The 95% confidence interval was not exceeded.
A linear regression analysis (least squares) was performed to assess a possible dose dependent increase of mutant frequencies. No significant dose dependent trend of the mutation frequency indicated by a probability value of <0.05 was determined in any of the experimental groups.
In the main experiment with and without S9 mix the range of the solvent controls was from 7.4 up to 24.7 mutants per 106 cells; the range of the groups treated with the test item was from 6.9 up to 25.9 mutants per 106 cells.
EMS (150 μg/mL) and DMBA (2.2 μg/mL) were used as positive controls and showed a distinct increase in induced mutant colonies. - Remarks on result:
- other: all strains/cell types tested
- Remarks:
- Migrated from field 'Test system'.
- Conclusions:
- Interpretation of results: negative
It can be stated that under the experimental conditions reported the test item did not induce gene mutations at the HPRT locus in V79 cells. Therefore, the test item is considered to be non-mutagenic in this HPRT assay. - Executive summary:
The study was performed to investigate the potential of test item to induce gene mutations at the HPRT locus in V79 cells of the Chinese hamster. The treatment period was 4 hours with and without metabolic activation. The maximum test item concentration of the pre-experiment (1642 μg/mL) was equal to a molar concentration of about 10 mM. The concentration range of the main experiment was limited by phase separation of the test item and cytotoxicity. No substantial and reproducible dose dependent increase of the mutation frequency was observed in the main experiment. Appropriate reference mutagens, used as positive controls, induced a distinct increase in mutant colonies and thus, showed the sensitivity of the test system and the activity of the metabolic activation system. In conclusion it can be stated that under the experimental conditions reported the test item did not induce gene mutations at the HPRT locus in V79 cells. Therefore, the test item is considered to be non-mutagenic in this HPRT assay.
- Endpoint:
- in vitro gene mutation study in bacteria
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- study well documented, meets generally accepted scientific principles, acceptable for assessment
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Deviations:
- no
- GLP compliance:
- no
- Type of assay:
- bacterial reverse mutation assay
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
- Species / strain / cell type:
- other: TA92
- Species / strain / cell type:
- other: TA94
- Species / strain / cell type:
- other: TA2637
- Metabolic activation:
- with and without
- Metabolic activation system:
- Aroclor 1254-induced rat liver S9 activation system
- Test concentrations with justification for top dose:
- Tested at 6 doses with a maximum of 5 mg/plate
- Vehicle / solvent:
- DMSO
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- Remarks:
- This conclusion can be drawn because of the validity of the Ames test.
- True negative controls:
- no
- Positive controls:
- yes
- Remarks:
- This conclusion can be drawn because of the validity of the Ames test.
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: According to Ames, McCann and Yamasaki (1975)
DURATION
- Preincubation period: Cells cultured overnight were pre-incubated with both the test sample and the S-9 mix for 20 min at 37 °C
- Exposure duration: 2 days - Evaluation criteria:
- The result was considered positive if the number of colonies found was twice the number in the control.
- Key result
- Species / strain:
- S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- not determined
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not specified
- Positive controls validity:
- valid
- Key result
- Species / strain:
- other: TA92
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- not determined
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not specified
- Positive controls validity:
- valid
- Key result
- Species / strain:
- other: TA94
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- not determined
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not specified
- Positive controls validity:
- valid
- Key result
- Species / strain:
- other: TA2637
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- not determined
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not specified
- Positive controls validity:
- valid
- Additional information on results:
- Up to 5 mg per plate no effects were seen.
- Remarks on result:
- other: all strains/cell types tested
- Remarks:
- Migrated from field 'Test system'.
- Conclusions:
- Interpretation of results
negative
In a mutagenesis study with the Salmonella strains TA92, TA1535, TA100, TA1537, TA94, TA98 and for some samples TA2637 with and without metabolic activation no mutagenic effects were observed. - Executive summary:
The test item was tested at 6 different doses by using Salmonella strains TA92, TA1535, TA100, TA1537, TA94, TA98 and for some samples TA2637 with and without S-9 liver microsome fraction. Each sample was tested at 6 doses; only maximum dose reported. The result was considered positive if the number of colonies found was twice the number in the control (exposed to the appropriate solvent or untreated). As vehicle DMSO was used. At 5 mg per plate no effects were observed.
- Endpoint:
- in vitro gene mutation study in bacteria
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 4 (not assignable)
- Rationale for reliability incl. deficiencies:
- abstract
- Remarks:
- Only one strain tested, only short abstract available
- Principles of method if other than guideline:
- Suspensions of bacterial cells are exposed to the test substance in the presence and in the absence of an exogenous metabolic activation system. Revertant colonies are counted and compared to the number of spontaneous revertant colonies.
- GLP compliance:
- no
- Type of assay:
- bacterial reverse mutation assay
- Species / strain / cell type:
- E. coli WP2 uvr A
- Metabolic activation:
- not specified
- Vehicle / solvent:
- DMSO
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in agar
- Key result
- Species / strain:
- E. coli WP2 uvr A
- Metabolic activation:
- not specified
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- not specified
- Vehicle controls validity:
- not specified
- Untreated negative controls validity:
- not specified
- Positive controls validity:
- not specified
- Additional information on results:
- No effects at 0.2-1.6 mg/plate.
- Remarks on result:
- other: all strains/cell types tested
- Remarks:
- Migrated from field 'Test system'.
- Conclusions:
- Interpretation of results (migrated information):
negative
The genotoxic potential of the test item was determined in a reverse mutation test in bacteria. No mutagenic potential was found. - Executive summary:
A mutagenic test was conducted with E.coli WP2 uvrA on 18 rec assay positive agents. DMSO was used as vehicle. No effects were observed at a concentration of 0.2 to 1.6 mg/plate.
Referenceopen allclose all
Experiment |
Preparation interval |
Test item concentration in µg/mL |
Mitototic indices in % of control |
incl. Gaps |
aberrant cells in % excl. Gaps |
with exchanges |
Exposure period 4 h without S9-Mix |
||||||
I |
22 h |
solvent control |
100.0 |
2.7 |
2.3 |
0.0 |
Positive control |
56.2 |
8.7 |
8.3 |
1.7 |
||
306.38 |
98.2 |
2.7 |
2.7 |
0.0 |
||
536.16 |
81.2 |
1.3 |
1.0 |
0.0 |
||
938.29 |
118.5 |
3.0 |
2.3 |
0.3 |
||
1642.00 |
71.0 |
1.7 |
1.7 |
0.0 |
||
Exposure period 22 h without S9 Mix |
||||||
IIA |
22 h |
Solvent control |
100.0 |
1.7 |
1.0 |
0.0 |
Positive control |
33.4 |
51.9 |
51.9 |
9.0 |
||
175.07 |
111.8 |
1.3 |
1.3 |
0.0 |
||
306.38 |
114.5 |
3.2 |
3.0 |
0.2 |
||
536.16 |
96.3 |
3.5 |
3.5 |
0.0 |
||
Exposure period 22 h without S9 Mix |
||||||
IIB |
22 h |
Solvent control |
100.0 |
1.3 |
1.3 |
0.0 |
Positive control |
53.7 |
20.3 |
20.0 |
5.7 |
||
402.22 |
81.1 |
1.0 |
0.7 |
0.0 |
||
536.29 |
86.3 |
3.0 |
3.0 |
0.3 |
||
670.36 |
67.7 |
2.0 |
1.7 |
0.0 |
||
Exposure period 4 h with S9 Mix |
||||||
I |
22 h |
Solvent control |
100.0 |
1.0 |
1.0 |
0.0 |
Positive control |
41.6 |
11.0 |
10.7 |
1.7 |
||
306.38 |
89.0 |
0.3 |
0.3 |
0.0 |
||
536.16 |
97.6 |
2.7 |
2.3 |
0.0 |
||
938.29 |
114.8 |
2.3 |
2.3 |
0.0 |
||
1642.00 |
117.7 |
2.0 |
1.7 |
0.3 | ||
IIA |
22 h |
Solvent control |
100.0 |
2.3 |
2.3 |
0.0 |
Positive control |
85.2 |
13.0 |
13.0 |
0.7 |
||
175.07 |
80.8 |
2.0 |
1.3 |
0.3 |
||
306.38 |
87.9 |
2.7 |
1.7 |
0.0 |
||
536.16 |
86.3 |
3.0 |
2.7 |
0.0 |
||
Conc. µg/mL |
PS |
S9 mix |
Relative cloning |
Relative cell |
Relative adjusted |
mutant colonies |
95 % confidence interval |
relative cloning efficiency I % |
relative cell density % |
relative adjusted cloning efficiency % |
mutant colonies |
95 % confidence interval |
|
Column |
1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
10 |
11 |
12 |
13 |
Exp I/4 h |
culture I |
culture II |
|||||||||||
Solvent control with DMSO |
- |
100.0 |
100.0 |
100.0 |
20.9 |
0.0-29.7 |
100.0 |
100.0 |
100.0 |
24.7 |
0.0-29.7 |
||
Positive control (EMS) |
150.0 |
- |
101.5 |
68.9 |
69.9 |
176.1 |
0.0-29.7 |
105.9 |
79.6 |
84.3 |
185.3 |
0.0-29.7 |
|
Test item |
25.6 |
- |
101.6 |
77.2 |
78.5 |
11.8 |
0.0-29.7 |
103.5 |
95.0 |
98.3 |
12.9 |
0.0-29.7 |
|
Test item |
51.3 |
- |
100.2 |
105.4 |
105.6 |
13.1 |
0.0-29.7 |
99.8 |
98.9 |
98.7 |
25.9 |
0.0-29.7 |
|
Test item |
102.5 |
- |
93.1 |
96.5 |
89.8 |
22.4 |
0.0-29.7 |
101.7 |
97.7 |
99.4 |
20.9 |
0.0-29.7 |
|
Test item |
205.0 |
- |
102.7 |
103.8 |
106.5 |
13.2 |
0.0-29.7 |
114.4 |
84.8 |
97.0 |
18.0 |
0.0-29.7 |
|
Test item |
410.0 |
PS |
- |
86.9 |
56.2 |
48.8 |
15.9 |
0.0-29.7 |
80.6 |
96.3 |
77.6 |
9.6 |
0.0-29.7 |
Test item |
820.0 |
PS |
- |
culture was not continued # |
culture was not continued # |
||||||||
Solvent control with DMSO |
+ |
100.0 |
100.0 |
100.0 |
7.4 |
0.0-27.7 |
100.0 |
100.0 |
100.0 |
13.7 |
0.0-27.7 |
||
Positive control (DMBA) |
2.2 |
+ |
66.8 |
171.4 |
114.5 |
151.3 |
0.0-27.7 |
79.4 |
104.8 |
83.2 |
140.4 |
0.0-27.7 |
|
Test item |
25.6 |
+ |
104.5 |
137.4 |
culture was not continued ## |
103.5 |
108.1 |
culture was not continued ## |
|||||
Test item |
51.3 |
+ |
102.1 |
104.1 |
106.3 |
23.6 |
0.0-27.7 |
104.9 |
109.3 |
114.7 |
7.5 |
0.0-27.7 |
|
Test item |
102.5 |
+ |
96.4 |
124.6 |
120.1 |
11.7 |
0.0-27.7 |
107.3 |
105.0 |
112.6 |
6.6 |
0.0-27.7 |
|
Test item |
205.0 |
+ |
103.6 |
145.5 |
150.8 |
24.5 |
0.0-27.7 |
98.2 |
101.2 |
99.4 |
14.5 |
0.0-27.7 |
|
Test item |
410.0 |
PS |
+ |
99.4 |
102.1 |
101.5 |
16.8 |
0.0-27.7 |
96.3 |
103.3 |
99.5 |
6.9 |
0.0-27.7 |
Test item |
820.0 |
PS |
+ |
98.8 |
122.6 |
121.2 |
21.6 |
0.0-27.7 |
103.3 |
96.9 |
100.1 |
11.6 |
0.0-27.7 |
PS= phase separation visible at the end of treatment |
|||||||||||||
# = culture was not continued due to exceedingly severe cytotoxic effects |
|||||||||||||
## = culture was not continued since a minimum of only four analysable concentrations is required |
|||||||||||||
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Additional information
A weight of evidence approach was performed to cover the endpoint in vitro gene mutation in bacteria.
Ames Test
Weight of evidence
Ames study of Ishidate, 1984 (Food Chem Toxicol 22 pg 623-636).
The test item was tested at 6 different doses by using Salmonella strains TA92, TA1535, TA100, TA1537, TA94, TA98 and for some samples TA2637 with and without S-9 liver microsome fraction. Each sample was tested at 6 doses; only maximum dose reported. The result was considered positive if the number of colonies found was twice the number in the control (exposed to the appropriate solvent or untreated). The test substance was tested in a purity of 99.9%. DMSO was used as vehicle at 5 mg per plate: No effects were observed
(Anti)Mutagenic acitivity test of Yoo, 1986 (J Osaka City Med Cent 34 (3-4) pg 1986).
A mutagenic test was conducted with E.coli WP2 uvrA on 18 rec assay positive agents. DMSO was used as vehicle. No effects were observed at concentrations from 0.2 to 1.6 mg/plate.
Conclusion: The mutagenic potential of the test item in bacteria was tested on 6 Salmonella strains and in E.coli WP2 uvrA. No mutagenic potential was found in these bacterias. In conclusion it can be stated the test substance is not mutagenic in bacteria.
Chromosome aberration test
The test item, dissolved in DMSO, was assessed for its potential to induce structural chromosomal aberrations in human lymphocytes in vitro in three independent experiments. In Experiment IIA in the absence of S9 mix no cytotoxicity was observed up to the highest evaluated concentration. Toxic effects, indicated by reduced mitotic indices were observed at higher concentrations. No evidence of an increase in polyploid metaphases was noticed after treatment with the test item as compared to the control cultures. In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce structural chromosomal aberrations in human lymphocytes in vitro. Therefore, it is considered to be non-clastogenic in this chromosome aberration test, when tested up to insoluble concentrations (phase separation) and in Experiment IIA up to the highest evaluable concentration.
HPRT test
The study was performed to investigate the potential of the test item to induce gene mutations at the HPRT locus in V79 cells of the Chinese hamster. The concentration range of the main experiment was limited by phase separation of the test item and cytotoxicity. No substantial and reproducible dose dependent increase of the mutation frequency was observed in the main experiment. In conclusion it can be stated that under the experimental conditions reported the test item did not induce gene mutations at the HPRT locus in V79 cells. Therefore, the test item is considered to be non-mutagenic in this HPRT assay.
Conclusion: 4 genetic toxicity studies were performed to determine the genetic toxicity potential of the test item. No genetic toxicity was observed in all studies.
Justification for selection of
genetic toxicity endpoint
Evaluation of genetic toxicity was done based on the GLP and
guideline in vitro studies (Ames, CA and HPRT)
Endpoint Conclusion: No adverse effect observed (negative)
Justification for classification or non-classification
Classification, Labelling, and Packaging Regulation (EC) No 1272/2008
The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. As a result the substance is not considered to be classified for genetic toxicity under Regulation (EC) No 1272/2008, as amended for the tenth time in Regulation (EU) No 2017/776.
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