Registration Dossier
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EC number: 270-115-0 | CAS number: 68411-30-3
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Endpoint summary
Administrative data
Key value for chemical safety assessment
Genetic toxicity in vitro
Description of key information
A series of in vitro studies were conducted to assess the potential mutagenicity of LAS. In the first study (Schoeberl 1993), a bacterial mutagenicity study (Ames test) was conducted using S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100, as well as TA1538 at test concentrations of 8, 40, 200, 1000 and 5000 µg/plate. All strains tested negative with and without S9 activation.
In the second test (Anon. 1995), the potential of LAS to cause mutations in mammalian cells was examined. Chinese Hamster Ovary (CHO) cells were exposed to concentrations of 0, 0.6, 1, 1.8, 3, and 6 µg/ml without S9, and 0, 6, 10, 18, 30, and 60 µg/ml with S9. The cells were then examined for cytogenicity and mutation frequency. Ethyl methane sulfonate and 3-(20-)methylcholanthrene were used as positive control substances. Preliminary tests show the test substance was cytotoxic at concentrations of 50 µg/ml or greater with metabolic activation, and 100 µg/ml or above without metabolic activation. There was no biologically significant increase in mutation frequency in the treated groups. Therefore, results show that LAS was not mutagenic to CHO cells both in the presence and absence of S9.
The third study (Murie and Innes 1997) examined the potential of LAS to cause chromosomal aberrations in mammalian cells. Chinese hamster ovary cells were exposed to concentrations of 2.5, 5, 10, 15, 20, 26, 33, and 39 µg/ml with S9, and 20, 39, 58, 78, 104, 130, and 156 µg/ml without S9. No biologically significant results were seen in treated cultures in the absence of metabolic activation. In the presence of metabolic activation the results were more equivocal. In the first of three tests, no cytotoxicity, and no increase in chromosome aberrations were observed at doses of 10 or 20 μg/ml and 100% cytotoxicity was observed at 39 μg/ml. In the second test, a steep cytotoxicity curve was observed between 10 and 20 μg/ml with a cell count of 68/90 % at the 10 μg/ml dose and no living cells remaining at the 20 μg/ml dose. An increased in aberration frequency could be observed at the 10 μg/ml dose. No increase in aberration frequency has been observed at lower doses which also did no show any cytotoxicity. To gain clarity on the positive result an additional test was conducted. Here, no cytotoxicity and no increase in chromosomal aberration frequency have been observed at the 10 μg/ml dose. At the 15 μg/ml dose the cell number was reduced to 25 % which is why this dose group cannot be evaluated due to excessive cytotoxicity. These results indicate that LAS is weakly clastogenic at cytotoxic concentrations but negative at concentrations below cytotoxic concentrations in this in vitro assay.
Link to relevant study records
- Endpoint:
- in vitro gene mutation study in bacteria
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: GLP laboratory study
- Justification for type of information:
- The test substance is a constituent of the reference substance, with similar alkyl chain range. Although the test substance does not contain sodium ions, at physiological pH, the test substance forms the same species as the dossier substance.
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
- Principles of method if other than guideline:
- Directive 84/449/EEC, B.14 Mutagenicity (Salmonella typhimurium - reverse mutation assay)" 1984; equivalent to OECD 471
- GLP compliance:
- yes
- Type of assay:
- bacterial reverse mutation assay
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
- Details on mammalian cell type (if applicable):
- also TA 1538
- Metabolic activation:
- with and without
- Metabolic activation system:
- Arochlor-induced S9 fraction
- Test concentrations with justification for top dose:
- 8, 40, 200, 1000 and 5000 ug/plate
- Vehicle / solvent:
- Water solution at 50 g/L
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Remarks:
- water
- True negative controls:
- yes
- Positive controls:
- yes
- Remarks:
- aminoanthracene
- Positive control substance:
- other: nitrofluorene, sodium azide and aminoacridine
- Species / strain:
- S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Remarks:
- with and without activation
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- TA 1538 also tested negative. During the pre-incubation test, signs of toxicity were noted at concentrations as low as 125 ug/plate. No precipitation of the product was observed at any concentration tested.
- Conclusions:
- LAS is not mutagenic in the Ames test.
- Executive summary:
A bacterial mutagenicity study (Ames test) was conducted on LAS and was found to be negative for mutagenicity.
- 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:
- May 16, 1995-June 30, 1995
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Comparable to guideline study.
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- mammalian cell gene mutation assay
- Species / strain / cell type:
- Chinese hamster Ovary (CHO)
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9 from aroclor 1254 induced rat liver
- Test concentrations with justification for top dose:
- 0, 0.6, 1, 1.8, 3, 6 ug/ml without S9
0, 6, 10, 18, 30, 60 ug/ml with S9 - Vehicle / solvent:
- None
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- no
- True negative controls:
- yes
- Remarks:
- H0 medium
- Positive controls:
- yes
- Positive control substance:
- other: ethyl methane sulfonate; 3-(20-)methylcholanthrene
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in medium
DURATION
- Preincubation period: 1 week
- Exposure duration: 4 hrs
- Expression time (cells in growth medium): 6 days at 37 degree C for cloning efficiency study, 9 days for mutation assay
STAIN (for cytogenetic assays): Giemsa
NUMBER OF REPLICATIONS: 2
- Evaluation criteria:
- A test substance was considered mutagenic if a statistically significant dose-related increase in mutant frequency was found in concentrations with greater than 20% survival rate. The mean mutant frequency must also be significantly above the maximum spontaneous mutant frequency.
- Statistics:
- Statistical significance was determined by the t-test.
- Species / strain:
- Chinese hamster Ovary (CHO)
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- preliminary test showed cytotoxicity at >= 50 ug/ml without S9, and >= 100 ug/ml with S9.
- Vehicle controls validity:
- not examined
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- COMPARISON WITH HISTORICAL CONTROL DATA: In both the studies with and without S9, the mutant frequencies in the treated groups were statistically significantly higher than in the concurrent negative controls. However, the mutant frequencies in the treated groups were not significantly increased when compared to historical negative controls. There was also no dose-response relationship. The increased mutant frequency in treated groups was therefore not considered to be biologically significant.
- Remarks on result:
- other: all strains/cell types tested
- Conclusions:
- Interpretation of results:
negative
The test substance is not mutagenic in either the presence or absence of metabolic activation. - Executive summary:
This study examined the potential of the test substance to cause mutations in mammalian cells. Chinese Hamster Ovary (CHO) cells were exposed to concentrations of 0, 0.6, 1, 1.8, 3, and 6 ug/ml without S9, and 0, 6, 10, 18, 30, and 60 ug/ml with S9. The cells were then examined for cytogenicity and mutation frequency. Ethyl methane sulfonate and 3-(20-)methylcholanthrene were used as positive control substances. Preliminary tests show the test substance was cytogenic at concentrations of 50 ug/ml or greater with metabolic activation, and 100 ug/ml or above without metabolic activation. There was no biologically significant increase in mutation frequency in the treated groups. The test substance is considered not mutagenic to CHO cells both in the presence and absence of S9.
- 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:
- May 25, 1995-November 23, 1995
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: GLP study done according to OECD guidelines. However, this study does not adequately address the results obtained at mildly cytotoxic concentrations.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- in vitro mammalian chromosome aberration test
- Species / strain / cell type:
- Chinese hamster Ovary (CHO)
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9 from Aroclor 1254 induced rat liver
- Test concentrations with justification for top dose:
- All concentrations in micrograms/ml
Test 1 with S9: 0.32, 0.63, 1.25, 2.5, 5, 10, 20, 39, 78
Test 1 without S9: 1.25, 2.5, 5, 10, 20, 39, 58,78, 156
Test 2 with S9: 2.5, 5, 10, 20, 26, 33, 39
Test 2 without S9: 20, 39, 58, 78, 130, 156
An additional test was done with S9 at the following dose levels:
2.5, 5, 7.5, 10, 15, 20, 25, and 30 ug/ml - Vehicle / solvent:
- None
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- no
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- other: methyl methanesulphonate, cyclophosphamide
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in medium
DURATION
- Exposure duration: 6 hrs with S9, 22 hrs without S9
- Expression time (cells in growth medium): 16-40 hrs with S9, 40 hrs without S9
- Selection time (if incubation with a selection agent): 2 hrs
- Fixation time (start of exposure up to fixation or harvest of cells): 24-48 hrs
SELECTION AGENT (mutation assays): Colcemid
NUMBER OF REPLICATIONS: 3
NUMBER OF CELLS EVALUATED: 100 metaphases
DETERMINATION OF CYTOTOXICITY
- Method: number of cells per culture
OTHER EXAMINATIONS:
- Determination of polyploidy: yes
- Determination of endoreplication: yes - Evaluation criteria:
- A dosage was considered toxic if cell count was less then 60% of cell cultures. A test substance was considered clastogenic if a single dose caused the percentage of aberrant cells to be consistently greater than the 99% confidence limits of negative controls and there was also an increase at another dose level.
- Statistics:
- 95% and 99% confidence limits
- Species / strain:
- Chinese hamster Ovary (CHO)
- Metabolic activation:
- with and without
- Genotoxicity:
- positive
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- >= 15 microgram/ml with S9, >=58 microgram/ml without S9
- Vehicle controls validity:
- not examined
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- In the absence of S9, only one culture (Test 2, 24 hr harvest, 20 ug/ml) showed a suspicious result. This single result was considered sporadic, as other cultures at this concentration, or at higher concentrations did not show a positive response. In Test 1, in the absence of S9, cytoxicity was seen at 78 micrograms/ml and above. In Test 2, in the absence of S9, cytoxicity was seen at concentrations of 58 micrograms/ml and above.
In Test 1, in the presence of S9, no positive results were seen at concentrations of up to 20 micrograms/ml. Metaphases could not be analyzed due to severe cytotoxicity at the 39 and 78 microgram/ml concentrations. In Test 2, in the presence of S9, one of the cultures at the 5 microgram/ml concentration gave a suspicious result, and both cultures at the 10 microgram/ml concentrations gave positive responses. Mild cytotoxity was also seen at the 10 microgram/ml concentration. At concentrations at and above 20 micrograms/ml, metaphases could not be analyzed due to severe cytotoxicity. No positive results were seen in the Test 2, 48 hr harvest cultures grown in the presence of S9, though moderate cytotoxicity was seen in one of the 20 microgram/ml cultures, and severe cytotoxicity was seen in all cultures above this concentration.
A third test was done in the presence of S9, which showed positive results at the 15 micrograms/ml concentration. However, this concentration was also moderately cytotoxic with only 26% of cells survival. However, due to the low survival of cells, these results are not definitive for determining clastogenicity. Higher concentrations were completely cytotoxic. An additional assessment was then performed at 10 micrograms/ml in the presence of S9, with negative results. - Remarks on result:
- other: all strains/cell types tested
- Conclusions:
- Interpretation of results:
negative without metabolic activation
positive with metabolic activation at cytotoxic concentrations or above
The test substance is not clastogenic in the absence of metabolic activation. The test substance is also not clastogenic in the presence of metabolic activation at non-cytotoxic concentrations. At cytotoxic concentrations, the test substance is weakly clastogenic. - Executive summary:
This study examined the potential of the test substance Marlon A 350 to cause chromosomal aberrations in mammalian cells. Chinese hamster ovary cells were exposed to concentrations of 0.32 to 78 ug/ml with S9, and 1.25 to 156 ug/ml without S9. Methyl methanesuflphonate and cyclophosphamide were used as positive controls. No biologically significant results were seen in treated cultures in the absence of metabolic activation. Positive responses were seen at cytotoxic concentrations in the presence of S9. Concentrations below the level of cytotoxicty with S9 did not show positive results. The test substance is not clastogenic in the absence of metabolic activation, or with metabolic activation below cytotoxic concentrations. These results indicate that LAS is weakly clastogenic at cytotoxic concentrations but negative at concentrations below cytotoxic concentrations
Referenceopen allclose all
Results of Test 1 ¿ Without S9 Mix
Concentration (ug/ml) | Absolute cloning efficiency (%) | Mutant frequency ( x 106) |
0 | 82 | 3 ± 2 |
0.6 | 86 | 7 ± 1 |
1 | 85 | 3 ± 2 |
1.8 | 78 | 5 ± 2 |
3 | 86 | 1 ± 1 |
6 | 83 | 0 ± 1 |
EMS | 83 | 277 ± 17 |
Results of Test 1 ¿ With S9 Mix
Concentration (ug/ml) | Absolute cloning efficiency (%) | Mutant frequency ( x 106) |
0 | 90 | 2 ± 1 |
6 | 88 | 1 ± 1 |
10 | 84 | 9 ± 4 |
18 | 78 | 5 ± 3 |
30 | 89 | 3 ± 2 |
60 | 89 | 7 ± 2 |
MCA | 81 | 91 ± 9 |
Results of Test 2 ¿ Without S9 Mix
Concentration (ug/ml) | Absolute cloning efficiency (%) | Mutant frequency ( x 106) |
0 | 96 | 1 ± 1 |
0.6 | 92 | 2 ± 3 |
1 | 95 | 1 ± 1 |
1.8 | 93 | 5 ± 2 |
3 | 90 | 2 ± 1 |
6 | 91 | 6 ± 6 |
EMS | 90 | 309 ± 20 |
Results of Test 2 ¿ With S9 Mix
Concentration (ug/ml) | Absolute cloning efficiency (%) | Mutant frequency ( x 106) |
0 | 90 | 2 ± 1 |
6 | 92 | 7 ± 3 |
10 | 88 | 9 ± 2 |
18 | 94 | 2 ± 1 |
30 | 93 | 2 ± 2 |
60 | 90 | 5 ± 1 |
MCA | 95 | 89 ± 6 |
Abbreviations used in tables:
T- Toxicity evident from morphological changes
TT- Toxicity evident from reduced cell count (<60% of vehicle)
TTT- Too toxic for metaphase assessment
Concentration (micrograms/ml) | Aberration Frequency (lesions/cell) | Aberrant Cell Frequency (% Including Gaps) | Aberrant Cell Frequency (% Excluding Gaps) | Cytotoxicity |
Ham's F10 medium | 0.01 | 1 | 0 | Nil |
Ham's F10 medium | 0.02 | 1 | 1 | Nil |
0.32 | - | - | - | Nil |
0.32 | - | - | - | Nil |
0.63 | - | - | - | Nil |
0.63 | - | - | - | Nil |
1.25 | - | - | - | Nil |
1.25 | - | - | - | Nil |
2.5 | 0.01 | 1 | 0 | Nil |
2.5 | 0.00 | 0 | 0 | Nil |
5 | 0.00 | 0 | 0 | Nil |
5 | 0.05 | 5 | 0 | Nil |
10 | 0.01 | 1 | 0 | Nil |
10 | 0.01 | 1 | 0 | Nil |
20 | 0.00 | 0 | 0 | Nil |
20 | 0.00 | 0 | 0 | Nil |
39 | - | - | - | TTT |
39 | - | - | - | TTT |
78 | - | - | - | TTT |
78 | - | - | - | TTT |
Cyclophosphamide (20 micrograms/ml) | 0.14 | 8 | 4 | - |
Cyclophosphamide (30 micrograms/ml) | 0.06 | 4 | 4 | - |
Cyclophosphamide (40 micrograms/ml) | 0.33 | 20 | 19 | - |
Test 1 ¿ Without S9 Mix, 24 hr Harvest
Concentration (micrograms/ml) | Aberration Frequency (lesions/cell) | Aberrant Cell Frequency (% Including Gaps) | Aberrant Cell Frequency (% Excluding Gaps) | Cytotoxicity |
Ham¿s F10 medium | 0.00 | 0 | 0 | Nil |
Ham¿s F10 medium | 0.00 | 0 | 0 | Nil |
1.25 | - | - | - | Nil |
1.25 | - | - | - | Nil |
2.5 | - | - | - | Nil |
2.5 | - | - | - | Nil |
5 | - | - | - | Nil |
5 | - | - | - | Nil |
10 | - | - | - | Nil |
10 | - | - | - | Nil |
20 | - | - | - | Nil |
20 | - | - | - | Nil |
39 | 0.01 | 1 | 0 | Nil |
39 | 0.00 | 0 | 0 | Nil |
58 | 0.01 | 1 | 0 | Nil |
58 | 0.00 | 0 | 0 | Nil |
78 | 0.00 | 0 | 0 | T |
78 | 0.00 | 0 | 0 | T |
156 | - | - | - | TTT |
156 | - | - | - | TTT |
Methyl methane-sulphonate (10 micrograms/ml) | 0.03 | 3 | 1 | - |
Cyclophosphamide (20 micrograms/ml) | 0.16 | 14 | 10 | - |
Test 2 ¿ With S9 Mix, 24 hr Harvest
Concentration (micrograms/ml) | Aberration Frequency (lesions/cell) | Aberrant Cell Frequency (% Including Gaps) | Aberrant Cell Frequency (% Excluding Gaps) | Cytotoxicity |
Ham¿s F-10 medium | 0.01 | 1 | 0 | Nil |
Ham¿s F-10 medium | 0.02 | 2 | 1 | Nil |
2.5 | 0.07 | 2 | 1 | Nil |
2.5 | 0.04 | 3 | 1 | Nil |
5 | 0.04 | 3 | 2 | Nil |
5 | 0.06 | 6 | 4 | Nil |
10 | 0.12 | 8 | 6 | T |
10 | 0.19 | 13 | 5 | T |
20 | - | - | - | TTT |
20 | - | - | - | TTT |
26 | - | - | - | TTT |
26 | - | - | - | TTT |
33 | - | - | - | TTT |
33 | - | - | - | TTT |
39 | - | - | - | TTT |
39 | - | - | - | TTT |
Cyclophosphamide (40 micrograms/ml) | 0.38 | 20 | 17 | - |
Cyclophosphamide (50 micrograms/ml) | 0.31 | 18 | 11 | - |
Test 2 ¿ With S9 Mix, 48 hr Harvest
Concentration (micrograms/ml) | Aberration Frequency (lesions/cell) | Aberrant Cell Frequency (% Including Gaps) | Aberrant Cell Frequency (% Excluding Gaps) | Cytotoxicity |
Ham¿s F-10 medium | 0.00 | 0 | 0 | Nil |
Ham¿s F-10 medium | 0.00 | 0 | 0 | Nil |
2.5 | 0.01 | 1 | 0 | Nil |
2.5 | 0.01 | 1 | 1 | Nil |
5 | 0.00 | 0 | 0 | Nil |
5 | 0.02 | 2 | 2 | Nil |
10 | 0.03 | 2 | 1 | Nil |
10 | 0.02 | 2 | 1 | TT |
20 | - | - | - | TTT |
20 | - | - | - | TTT |
26 | - | - | - | TTT |
26 | - | - | - | TTT |
33 | - | - | - | TTT |
33 | - | - | - | TTT |
39 | - | - | - | TTT |
39 | - | - | - | TTT |
Cyclophosphamide (40 micrograms/ml) | 0.03 | 3 | 2 | - |
Cyclophosphamide (50 micrograms/ml) | 0.10 | 8 | 7 | - |
Test 2 ¿ Without S9 Mix, 24 hr Harvest
Concentration (micrograms/ml) | Aberration Frequency (lesions/cell) | Aberrant Cell Frequency (% Including Gaps) | Aberrant Cell Frequency (% Excluding Gaps) | Cytotoxicity |
Ham¿s F-10 medium | 0.02 | 2 | 2 | Nil |
Ham¿s F-10 medium | 0.03 | 3 | 0 | Nil |
20 | 0.02 | 2 | 0 | Nil |
20 | 0.05 | 5 | 3 | Nil |
39 | 0.02 | 2 | 1 | Nil |
39 | 0.04 | 4 | 0 | Nil |
58 | 0.01 | 1 | 1 | Nil |
58 | 0.06 | 6 | 1 | Nil |
78 | - | - | - | TTT |
78 | - | - | - | TTT |
104 | - | - | - | TTT |
104 | - | - | - | TTT |
130 | - | - | - | TTT |
130 | - | - | - | TTT |
156 | - | - | - | TTT |
156 | - | - | - | TTT |
Methyl methane-sulphonate (10 micrograms/ml) | 0.30 | 21 | 14 | - |
Methyl methane-sulphonate (20 micrograms/ml) | 0.71 | 33 | 28 | - |
Test 2 ¿ Without S9 Mix, 48 hr Harvest
Concentration (micrograms/ml) | Aberration Frequency (lesions/cell) | Aberrant Cell Frequency (% Including Gaps) | Aberrant Cell Frequency (% Excluding Gaps) | Cytotoxicity |
Ham¿s F-10 medium | 0.01 | 1 | 1 | Nil |
Ham¿s F-10 medium | 0.00 | 0 | 0 | Nil |
20 | 0.00 | 0 | 0 | Nil |
20 | 0.00 | 0 | 0 | Nil |
39 | 0.01 | 1 | 1 | Nil |
39 | 0.00 | 0 | 0 | Nil |
58 | 0.00 | 0 | 0 | T |
58 | 0.01 | 1 | 0 | T |
78 | - | - | - | TTT |
78 | - | - | - | TTT |
104 | - | - | - | TTT |
104 | - | - | - | TTT |
130 | - | - | - | TTT |
130 | - | - | - | TTT |
156 | - | - | - | TTT |
156 | - | - | - | TTT |
Methyl methane-sulphonate (20 micrograms/ml) | 0.21 | 11 | 8 | - |
Methyl methane- sulphonate (40 micrograms/ml) | 3.20 | 60 | 60 | - |
Test 3 ¿ With S9 Mix, 24 hr Harvest
Concentration (micrograms/ml) | Aberration Frequency (lesions/cell) | Aberrant Cell Frequency (% Including Gaps) | Aberrant Cell Frequency (% Excluding Gaps) | Cytoxicity |
Ham¿s F-10 medium | 0.04 | 4 | 0 | Nil |
Ham¿s F-10 medium | 0.04 | 4 | 0 | Nil |
2.5 | - | - | - | Nil |
2.5 | - | - | - | Nil |
5 | - | - | - | Nil |
5 | - | - | - | Nil |
7.5 | - | - | - | Nil |
7.5 | - | - | - | Nil |
10 | - | - | - | Nil |
10 | - | - | - | Nil |
15 | 0.20 | 12 | 8 | TT |
15 | 0.18 | 12 | 6 | TT |
20 | - | - | - | TTT |
20 | - | - | - | TTT |
25 | - | - | - | TTT |
25 | - | - | - | TTT |
30 | - | - | - | TTT |
30 | - | - | - | TTT |
Cyclophosphamide (30 micrograms/ml) | 0.24 | 14 | 12 | - |
Cyclophosphamide (40 micrograms/ml) | 0.32 | 17 | 11 | - |
Test 3 - see tables below
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Genetic toxicity in vivo
Description of key information
A robust in vivo mammalian micronucleus study is available on the structurally related substance LAS Acid (CAS#85536-14-7, Benzenesulfonic acid, C10-13-sec alkyl derivatives). In this study (Fedtke 1991), 40 male and 40 female mice were given a single oral dose by gavage of 1122 mg/kg LAS Acid (read across) and evaluated for chromosome aberrations. Only a single dose has been evaluated which was in the range of the acute oral LD50 value for LAS Acid in rats (LD50 = 1470 mg/kg). Furthermore, slight cytotoxicity has been observed after 48 hours. No statistically significant or biologically relevant increases in the number of polychromatic erythrocytes with micronuclei were observed; therefore the test material is considered negative for cytogenicity.
In an in vivo mammalian chromosome aberration study on LAS by Inoue et al. 1976, groups of male mice were given doses of 200, 400, or 800 mg/kg of LAS. This is about half the acute oral LD50 of 1655 mg/kg, as cited by the authors of the study. Mice were sacrificed at 6, 24, and 48 hrs, three of the mice from each dosage group were sacrificed, and bone marrow cells from the femurs collected and examined for chromosome aberrations. In addition, one group of mice was exposed daily for 5 consecutive days. Additional groups of mice were exposed to commercial detergents containing 19% or 17.1% of LAS. None of the treatment groups showed any significant increase in chromosome aberrations as compared to negative controls while the positive control, Mitomycin C, clearly showed an increase in chromosomal aberration frequency. Therefore, LAS was not considered clastogenic in this assay.
A second chromosomal aberration study was conducted by Masubuchi et al. 1976. In this study, groups of five male mice or five male rats were fed diets containing 0.9% LAS for 9 months (1125 mg/kg bw/d for mice, 405 mg/kg bw/d for rats). At the end of this period, the animals were sacrificed, and the bone marrow cells examined for chromosome aberrations. No increase in chromosome aberrations was seen as compared to negative controls in either species but no positive controls have been included in this assay. LAS was not clastogenic in this assay.
These same authors (Masubuchi et al. 1976) also conducted a dominant lethal study in mice. A group of seven male mice were fed a diet containing 0.6% LAS for 9 months (750 mg/kg bw/d). At the end of this period they were each mated with two untreated females. Females were sacrificed on day 13 of gestation for examination of ovaries and uteri. No evidence of dominant lethal mutations was observed as compared to the controls.Link to relevant study records
- Endpoint:
- in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- read-across based on grouping of substances (category approach)
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: GLP laboratory study
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- micronucleus assay
- Species:
- mouse
- Strain:
- NMRI
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- Strain NMRI. Animals were approximately 22-26 g (male) and 20-25 g (female) and acclimated for 1 week to the test conditions (20 =/- 3 degrees C, 30-70% relative humidity, 12 hour light/dark cycle). Food was given daily and water was ad libitum. All animals were healthy at the time of test initiation.
- Route of administration:
- oral: gavage
- Vehicle:
- NaCl
- Duration of treatment / exposure:
- 72 hours
- Frequency of treatment:
- single dose
- Remarks:
- Doses / Concentrations:
1122 mg/kg
Basis:
actual ingested - No. of animals per sex per dose:
- 40 males and 40 females per dose
- Control animals:
- yes
- Positive control(s):
- Endoxan (cyclophosphamid)
- Tissues and cell types examined:
- Cells were taken from the thigh.
- Details of tissue and slide preparation:
- Cells were mixed with cattle serum and suspended, then centrifuged. The sediment was then resuspended. The suspension was seperated in a cellulose chromatography column. This was centrifuged, and mixed with fetal calf serum and EDTA. This was air-dried for 24 hrs and stained with Giemsa.
- Evaluation criteria:
- number of polychromatid erythrocytes (PCE)
ratio of PCE to normochromatid erythrocytes (NCE)
number of cells with micronucleus - Sex:
- male/female
- Genotoxicity:
- negative
- Toxicity:
- no effects
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Conclusions:
- Interpretation of results: negative
- Executive summary:
No significant increases in the number of polychromatic erythrocytes with micronuclei were observed.
- Endpoint:
- in vivo mammalian germ cell study: cytogenicity / chromosome aberration
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Well-documented publication.
- Qualifier:
- no guideline followed
- Principles of method if other than guideline:
- Groups of male mice were given doses of 200, 400, or 800 mg/kg of Benzenesulfonic acid, C10-14-alkyl derivs., sodium salts. At 6, 24, and 48 hrs, 3 of the mice from each dosage group were sacrificed. The bone marrow cells from the femurs were collected and examined for chromosome aberrations. In addition, one group of mice was exposed daily for 5 consecutive days. Additional groups of mice were exposed to commerical detergents containing 19% or 17.1% of the test substance. Mitomycin C was used as a positive control.
- GLP compliance:
- no
- Type of assay:
- chromosome aberration assay
- Species:
- mouse
- Strain:
- other: ICR/JCL
- Sex:
- male
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: CLEA Japan Inc.
- Age at study initiation: 9-11 weeks
- Housing: individually in plastic cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): tap water ad libitum
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 +/- 2
- Humidity (%): 55 +/- 5
- Photoperiod (hrs dark / hrs light): 12/12 hrs - Route of administration:
- oral: gavage
- Vehicle:
- - Vehicle(s)/solvent(s) used: distilled water
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS: Samples were diluted to make a 5 ml/kg dose volume
- Duration of treatment / exposure:
- Single treatment, except for one group which was given 5 consecutive daily exposures to the test substance
- Frequency of treatment:
- once daily
- Remarks:
- Doses / Concentrations:
200, 400, 800 mg/kg
Basis:
nominal conc.
test substance - Remarks:
- Doses / Concentrations:
800, 1600, 3200 mg/kg
Basis:
nominal conc.
commercial detergent containing 19% test substance - Remarks:
- Doses / Concentrations:
1000, 2000, 4000 mg/kg
Basis:
nominal conc.
commercial detergent containing 17.1% test substance - No. of animals per sex per dose:
- 9
- Control animals:
- yes, concurrent vehicle
- Positive control(s):
- mitomycin C1
- Route of administration: intraperitoneally
- Doses / concentrations: 5 mg/kg - Tissues and cell types examined:
- bone marrow cells from femurs
- Details of tissue and slide preparation:
- CRITERIA FOR DOSE SELECTION:
TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): 3 animals were sacrificed at 6, 24, and 48 hrs after treatment. One group was exposed daily for 5 days prior to sacrifice.
DETAILS OF SLIDE PREPARATION: Cells were placed in Hank's solution, then centrifuged at 1000 rpm for 5 min. Supernatant was discarded, and 5 ml of 0.075 M KCl was added. The mixture then stood for 5 min., then was stirred and centrifuged again. This was repeated several times, before finally placing one or two drops on the slides, and staining with 2% Giemsa solution. - Evaluation criteria:
- number of cells with chromatid and chromosome gaps, number of cells with aberrations
50 metaphases per animal (150 total) were imaged at each time point. - Sex:
- male
- Genotoxicity:
- negative
- Toxicity:
- not examined
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- No significant differences in the incidence of chromosomal aberrations were observed in any test substance treatment group relative to the controls.
- Conclusions:
- Interpretation of results: negative
The test substance is not clastogenic. - Executive summary:
Groups of male mice were given doses of 200, 400, or 800 mg/kg of Benzenesulfonic acid, C10-14-alkyl derivs., sodium salts. At 6, 24, and 48 hrs, 3 of the mice from each dosage group were sacrificed. The bone marrow cells from the femurs were collected and examined for chromosome aberrations. In addition, one group of mice was exposed daily for 5 consecutive days. Additional groups of mice were exposed to commerical detergents containing 19% or 17.1% of the test substance. Mitomycin C was used as a positive control. None of the treatment groups showed any significant increase in chromosome aberrations as compared to negative controls. The test substance in not clastogenic.
- Endpoint:
- in vivo mammalian germ cell study: cytogenicity / chromosome aberration
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Well-documented journal article.
- Qualifier:
- no guideline followed
- Principles of method if other than guideline:
- A group of 5 male mice was fed a diet containing 0.9% test substance for 9 months. At the end of this period, the animals were sacrificed, and the bone marrow cells examined for chromosome aberrations.
- GLP compliance:
- no
- Type of assay:
- mammalian germ cell cytogenetic assay
- Species:
- mouse
- Strain:
- other: JCL-ICR
- Sex:
- male
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: CLEA Japan, Inc.
- Age at study initiation: 4 weeks
- Housing: individually, except during breeding
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period:
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 25 +/- 1
- Humidity (%): 55 +/- 5
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: feed
- Details on exposure:
- DIET PREPARATION
- Mixing appropriate amounts with (Type of food): feed powder CE-2 - Duration of treatment / exposure:
- 9 months
- Frequency of treatment:
- daily
- Remarks:
- Doses / Concentrations:
0.9%, 1170 mg/kg bw d
Basis:
nominal in diet - No. of animals per sex per dose:
- 5
- Control animals:
- yes
- Tissues and cell types examined:
- femur bone marrow cells
- Details of tissue and slide preparation:
- Animals were sacrificed by administration of 1 ml/kg of 1% colchine solution. Femurs were then removed, and bone marrow cells washed into centrifuge tubes. The cells were then treated with 0.075 M KCl solution at 37 degree C for 15 min, and then fixed with an acetic acid 1: ethanol 3 solution. Samples were then flame dried and treated with Giemsa.
- Evaluation criteria:
- Presence and absence of chromosomal aberrations. 50 metaphases per individual.
- Statistics:
- Rohrborn's method.
- Sex:
- male
- Genotoxicity:
- negative
- Negative controls validity:
- valid
- Additional information on results:
- No increase in chromosome aberrations was noted.
- Conclusions:
- Interpretation of results: negative
The test substance is not clastogenic. - Executive summary:
A group of 5 male mice was fed a diet containing 0.9% test substance for 9 months. At the end of this period, the animals were sacrificed, and the bone marrow cells examined for chromosome aberrations. No increase in chromosome aberrations was seen as compared to controls. The test substance is not clastogenic.
- Endpoint:
- in vivo mammalian germ cell study: cytogenicity / chromosome aberration
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Well-documented journal article.
- Qualifier:
- no guideline followed
- Principles of method if other than guideline:
- Groups of 5 male rats were fed a diet containing 0.9% test substance for 9 months. At the end of this period, the animals were sacrificed, and the bone marrow cells examined for chromosome aberrations.
- GLP compliance:
- no
- Type of assay:
- mammalian germ cell cytogenetic assay
- Species:
- rat
- Strain:
- other: Wistar and SD
- Sex:
- male
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: CLEA Japan, Inc.
- Age at study initiation: 4 weeks
- Housing: individually
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period:
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 25 +/- 1
- Humidity (%): 55 +/- 5
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: feed
- Details on exposure:
- DIET PREPARATION
- Mixing appropriate amounts with (Type of food): feed powder CE-2 - Duration of treatment / exposure:
- 9 months
- Frequency of treatment:
- daily
- Remarks:
- Doses / Concentrations:
0.9%, 450 mg/kg bw d
Basis:
nominal in diet - No. of animals per sex per dose:
- 10
- Control animals:
- yes
- Tissues and cell types examined:
- femur bone marrow cells
- Details of tissue and slide preparation:
- Animals were sacrificed by administration of 1 ml/kg of 1% colchine solution. Femurs were then removed, and bone marrow cells washed into centrifuge tubes. The cells were then treated with 0.075 M KCl solution at 37 degree C for 15 min, and then fixed with an acetic acid 1: ethanol 3 solution. Samples were then flame dried and treated with Giemsa.
- Evaluation criteria:
- Presence and absence of chromosomal aberrations. 50 metaphases per individual.
- Statistics:
- Rohrborn's method.
- Sex:
- male
- Genotoxicity:
- negative
- Negative controls validity:
- valid
- Additional information on results:
- No increase in chromosome aberrations was noted.
- Conclusions:
- Interpretation of results: negative
The test substance is not clastogenic. - Executive summary:
Groups of 5 male rats were fed a diet containing 0.9% test substance for 9 months. At the end of this period, the animals were sacrificed, and the bone marrow cells examined for chromosome aberrations. No increase in chromosome aberrations was seen as compared to controls. The test substance is not clastogenic.
- Endpoint:
- in vivo mammalian germ cell study: cytogenicity / chromosome aberration
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Well-documented journal article.
- Qualifier:
- no guideline followed
- Principles of method if other than guideline:
- A group of 7 male mice was fed a diet containing 0.6% test substance for 9 months. At the end of this period, the animals were each mated with two untreated females. On day 13 of pregnancy, the females were sacrificed, and the ovaries and uteri were examined.
- GLP compliance:
- no
- Type of assay:
- rodent dominant lethal assay
- Species:
- mouse
- Strain:
- other: JCL-ICR
- Sex:
- male
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: CLEA Japan, Inc.
- Age at study initiation: 4 weeks
- Housing: individually, except during breeding
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 25 +/- 1
- Humidity (%): 55 +/- 5
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: feed
- Details on exposure:
- DIET PREPARATION
- Mixing appropriate amounts with (Type of food): feed powder CE-2 - Duration of treatment / exposure:
- 9 months
- Frequency of treatment:
- daily
- Remarks:
- Doses / Concentrations:
0.6%, 780 mg/kg bw d
Basis:
nominal in diet - No. of animals per sex per dose:
- 7
- Control animals:
- yes
- Statistics:
- Rohrborn's method.
- Sex:
- male
- Genotoxicity:
- negative
- Negative controls validity:
- valid
- Additional information on results:
- There were no significant differences in fertility, the mortality of ova and embryos, the number of surviving fetuses, or the index of dominant lethal induction between the experimental groups and the control group.
- Conclusions:
- Interpretation of results: negative
The test substance did not cause genetic disorders in mice. - Executive summary:
A group of 7 male mice was fed a diet containing 0.6% test substance for 9 months. At the end of this period, the animals were each mated with two untreated females. On day 13 of pregnancy, the females were sacrificed, and the ovaries and uteri were examined. No increase in dominant lethal induction was seen as compared to controls. The test substance does not cause genetic disorders.
Referenceopen allclose all
No significant increases in the number of polychromatic erythrocytes with micronuclei were observed.
Chromosome Aberrations
| Total number of cells having aberrations and occurrence (%) | |||
| 6 hrs | 24 hrs | 48 hrs | 5 days |
200 mg/kg | 0 (0) | 0 (0) | 0 (0) | 1 (0.7) |
400 mg/kg | 1 (0.7) | 0 (0) | 0 (0) | 0 (0) |
800 mg/kg | 0 (0) | 0 (0) | 0 (0) | 0 (0) |
800 mg/kg of 17.1% detergent | 0 (0) | 0 (0) | 0 (0) | - |
1600 mg/kg of 17.1% detergent | 0 (0) | 1 (0.7) | 0 (0) | - |
3200 mg/kg of 17.1% detergent | 2 (1.3) | 2 (1.3) | 0 (0) | - |
1000 mg/kg of 19% detergent | - | 0 (0) | - | - |
2000 mg/kg of 19% detergent | - | 0 (0) | - | - |
4000 mg/kg of 19% detergent | - | 0 (0) | - | - |
Mitomycin C | 16 (10.7) | 53 (353) | 13 (8.7) | 112 (74.7) |
Distilled water | 0 (0) | 0 (0) | 0 (0) | 0 (0) |
untreated | 0 (0) | 0 (0) | 1 (0.7) | 0 (0) |
Chromosome Aberrations
| 0.9% in Diet | Control |
No. of cells with chromatid breaks | 1 | 2 |
No. of cells with isochromatid breaks | 1 | 0 |
No. of cells with chromatid gaps | 4 | 5 |
No. of cells with isochromatid gaps | 0 | 0 |
No. of cells with other aberrations | 0 | 0 |
Chromosome Aberrations
| 0.9% in Diet ¿ Wister Rats | 0.9% in Diet ¿ SD Rats | Control | Control |
No. of cells with chromatid breaks | 0 | 0 | 1 | 0 |
No. of cells with isochromatid breaks | 0 | 0 | 0 | 0 |
No. of cells with chromatid gaps | 3 | 4 | 3 | 4 |
No. of cells with isochromatid gaps | 0 | 0 | 0 | 0 |
No. of cells with other aberrations | 0 | 0 | 0 | 0 |
Dominant Lethal Assay Results
| 0.6% in Diet | Control |
Number of mating females | 14 | 18 |
Number pregnant | 11 | 12 |
No. with dead embryos | 6 | 10 |
Dead embryos per pregnant female | 54.6% | 83.3% |
No. of corpora lutea | 156 | 161 |
Corpora lutea per pregnant female | 14.2 | 13.4 |
No. of implants | 148 | 156 |
Implants per pregnant female | 13.5 | 13.0 |
Implants per corpora lutea | 94.9 | 96.9 |
No. of live fetuses | 142 | 143 |
Live fetuses per pregnant female | 12.9 | 11.9 |
Live fetuses per corpora lutea | 91.0 | 88.8 |
Live fetuses per total implants | 96.0 | 91.7 |
No. of early dead fetuses | 4 | 12 |
No. of late dead fetuses | 2 | 1 |
% of dominant lethals | -4.67 | - |
% of dominant lethals | -8.33 | - |
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Additional information
LAS was consistently found not to cause induction of gene mutations in the Ames bacterial reverse mutation assay as well as in the OECD 476 in vitro mammalian cell gene mutation test. LAS was found to be positive at cytotoxic concentrations with metabolic activation, but negative at non-cytotoxic concentrations with metabolic activation and negative without metabolic activation, when tested in an in vitro chromosome aberration assay in CHO cells. When tested in a battery of in vivo genotoxicity studies, LAS was consistently found not to cause any mutagenic or clastogenic responses. An additional study conducted on the structural analogue LAS Acid further supports that LAS is not expected to be mutagenic or clastogenic. The positive result in the in vitro chromosome aberration study using a rodent cell line (CHO cells) derived from cancer tissues that is lacking proper cell cycle control has to be seen in the context of the extensive in vivo data. In vivo studies do assess genotoxicity under more realistic conditions, including eADME and metabolic activation.
Justification for selection of genetic toxicity endpoint
In vivo experimental studies, consistent with in vitro mutagenicity studies, considered more reliable than in vitro cytogenic study.
Justification for classification or non-classification
Based on the consistent absence of mutagenic or clastogenic responses in in vivo studies using LAS or the structural analogue LAS Acid, LAS is not expected to be mutagenic or clastogenic.
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