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EC number: 214-290-3 | CAS number: 1119-94-4
- 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
Description of key information
Using Derek Nexus v5.0, the skin sensitising potential of the test item was estimated to be equivocal (reference 7.4.1-1). However, basaed on the results of an in chemico/in vitro test strategy the test item is not peptide reactive (DPRA,OECD TG 442C, reference 7.4.1-2) and does not activate keratinocytes (Keratinosens, OECD TG 442D, reference 7.4.1-3). Therefore, the substance is not predicted to be a skin sensitizer.
Key value for chemical safety assessment
Skin sensitisation
Link to relevant study records
- Endpoint:
- skin sensitisation, other
- Remarks:
- in silico
- Type of information:
- (Q)SAR
- Adequacy of study:
- weight of evidence
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- results derived from a valid (Q)SAR model and falling into its applicability domain, with adequate and reliable documentation / justification
- Justification for type of information:
- Please refer to the QMRF and QPRF files provided under the section attached justification.
- Qualifier:
- no guideline available
- Principles of method if other than guideline:
- Estimates the skin sensitising properties of chemicals using structural alert relationships.
- GLP compliance:
- no
- Specific details on test material used for the study:
- SMILES: [Br-].CCCCCCCCCCCC[N+](C)(C)C
- Key result
- Parameter:
- other: alerts
- Value:
- 1
- Remarks on result:
- other: Skin sensitisation in mammal is equivocal. Alert matched: 436 Quaternary ammonium salt
- Remarks:
- QSAR predicted value. The substance is within the applicability domain of the model.
- Interpretation of results:
- other: Derek result: equivocal
- Conclusions:
- Using Derek Nexus v5.0, the skin sensitising potential of the test item was estimated to be equivocal. The substance is within the applicability domain of the model. Thus the estimation can be regarded as accurate.
- Executive summary:
The skin sensitising properties were estimated using Derek Nexus v5.0. The skin sensitising potential of the test item was estimated to be equivocal based on the described QSAR method (Derek, 2017).
The adequacy of a prediction depends on the following conditions:
a) the (Q)SAR model is scientifically valid: the scientific validity is established according to the OECD principles for (Q)SAR validation;
b) the (Q)SAR model is applicable to the query chemical: a (Q)SAR is applicable if the query chemical falls within the defined applicability domain of the model;
c) the (Q)SAR result is reliable: a valid (Q)SAR that is applied to a chemical falling within its applicability domain provides a reliable result;
d) the (Q)SAR model is relevant for the regulatory purpose.
For assessment and justification of these 4 requirements the QMRF and QPRF files were developed and attached to this study record.
Description of the prediction Model
The prediction model was descripted using the harmonised template for summarising and reporting key information on (Q)SAR models. For more details please refer to the attached QSAR Model Reporting Format (QMRF) file.
Assessment of estimation domain
The assessment of the estimation domain was documented in the QSAR Prediction Reporting Format file (QPRF). Please refer to the attached document for the details of the prediction and the assessment of the estimation domain.
- Endpoint:
- skin sensitisation: in chemico
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 2018-01-24 to 2018-02-08
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 442C (In Chemico Skin Sensitisation: Direct Peptide Reactivity Assay (DPRA))
- Version / remarks:
- 04 February 2015
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: Direct Peptide Reactivity Assay (DPRA) for Skin Sensitization Testing, DB-ALM Protocol n°154, January 12, 2013
- Version / remarks:
- 12 January 2013
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany
- Type of study:
- other: (in chemico) reactivity against synthetic peptides with a thiol or amino group
- Details on the study design:
- The in chemico direct peptide reactivity assay (DPRA) enables detection of the sensitising potential of a test item by quantifying the reactivity of test chemicals towards synthetic peptides containing either lysine or cysteine.
- Positive control results:
- The 100 mM stock solution of the positive control (cinnamic aldehyde) showed high reactivity towards the synthetic peptides. The mean depletion of both peptides was 67.89%.
- Key result
- Run / experiment:
- other: cysteine run
- Parameter:
- other: mean peptide depletion [%]
- Value:
- 0.1
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Key result
- Run / experiment:
- other: lysine run
- Parameter:
- other: mean peptide depletion [%]
- Value:
- 0
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Other effects / acceptance of results:
- Acceptance Criteria
The run meets the acceptance criteria if:
- the standard calibration curve has a r² > 0.99,
- the mean percent peptide depletion (PPD) value of the three replicates for the positive control is
between 60.8% and 100% for the cysteine peptide and the maximum standard deviation (SD) for the
positive control replicates is < 14.9%,
- the mean percent peptide depletion (PPD) value of the three replicates for the positive control is
between 40.2% and 69.0% for the lysine peptide and the maximum SD for the positive control
replicates is < 11.6%,
- the mean peptide concentration of the three reference controls A replicates is 0.50 ± 0.05 mM,
- the coefficient of variation (CV) of peptide peak areas for the six reference control B replicates and three reference control C replicates in acetonitrile is < 15.0%.
The results of the test item meet the acceptance criteria if:
- the maximum standard deviation (SD) for the test chemical replicates is < 14.9% for the cysteine
percent depletion (PPD),
- the maximum standard deviation (SD) for the test chemical replicates is < 11.6% for the lysine
percent depletion (PPD),
- the mean peptide concentration of the three reference controls C replicates in the appropriate solvent is 0.50 ± 0.05 mM.
Both peptide runs and the test item results met the acceptance criteria of the test. - Conclusions:
- In this study under the given conditions the test item showed minimal reactivity towards both peptides. The test item might be considered as “non-sensitiser”.
The data generated with this method may be not sufficient to conclude on the absence of skin sensitisation potential of chemicals and should be considered in the context of integrated approach such as IATA. - Executive summary:
In the present study dodecyltrimethylammonium bromide was dissolved in acetonitrile, based on the results of the pre-experiments. Based on a molecular weight of 308.34 g/mol a 100 mM stock solution was prepared. The test item solutions were tested by incubating the samples with the peptides containing either cysteine or lysine for 24 ± 2 h at 25 ± 2.5 °C. Subsequently samples were analysed by HPLC.
For the 100 mM solution of the test item no turbidity or precipitation was observed when diluted with the cysteine peptide solution. After the 24 h±2 h incubation period but prior to the HPLC analysis samples were inspected for precipitation, turbidity or phase separation. No precipitation, turbidity or phase separation was observed for any of the samples.
For the 100 mM solution of the test item no turbidity or precipitation was observed when diluted with the lysine peptide solution. After the 24 h±2 h incubation period but prior to the HPLC analysis samples were inspected for precipitation, turbidity or phase separation. Phase separation was observed for the co-elution control of the positive control. No precipitation, turbidity or phase separation was observed for the samples of the test item. Samples were not centrifuged prior to the HPLC analysis.
Since the acceptance criteria for the depletion range of the positive control were fulfilled, the observed phase separation was regarded as insignificant.
No co-elution of test item with the peptide peaks was observed. Sensitising potential of the test item was predicted from the mean peptide depletion of both analysed peptides (cysteine and lysine) by comparing the peptide concentration of the test item treated samples to the corresponding reference control C (RC C acetonitrile).
The 100 mM stock solution of the test item showed minimal reactivity towards the synthetic peptides. The mean depletion of both peptides was ≤ 6.38% (0.05%). Based on the prediction model 1 the test item can be considered as non-sensitiser.
The 100 mM stock solution of the positive control (cinnamic aldehyde) showed high reactivity towards the synthetic peptides. The mean depletion of both peptides was 67.89%.
The controls confirmed the validity of the study for both the cysteine and lysine run.
- Endpoint:
- skin sensitisation: in vitro
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 2018-03-15 to 2018-03-26
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 442D (In Vitro Skin Sensitisation: ARE-Nrf2 Luciferase Test Method)
- Version / remarks:
- 04 February 2015
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: KeratinoSens™, EURL ECVAM DB-ALM Protocol No. 155, July 1st, 2015
- Version / remarks:
- 01 July 2015
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany
- Type of study:
- activation of keratinocytes
- Details on the study design:
- The in vitro KeratinoSens™ assay enables detection of the sensitising potential of a test item by addressing the second molecular key event of the adverse outcome pathway (AOP), namely activation of keratinocytes, by quantifying the luciferase activity in the transgenic cell line KeratinoSens™. The luciferase activity, assessed by luminescence measurement, compared to the respective solvent controls is used to support discrimination between skin sensitisers and non-sensitisers.
- Positive control results:
- The luciferase activity induced by the positive control at a concentration of 64 µM was between 2 and 8 (5.80 (experiment 1); 3.67 (experiment 2)).
- Key result
- Run / experiment:
- other: 1
- Parameter:
- other: luciferase activity
- Value:
- 1.34
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: Concentration: 15.63µM
- Key result
- Run / experiment:
- other: 1
- Parameter:
- other: cell viability [%]
- Value:
- 12.9
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Key result
- Run / experiment:
- other: 1
- Parameter:
- other: EC1.5 [µM]
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- not determinable
- Key result
- Run / experiment:
- other: 2
- Parameter:
- other: luciferase activity
- Value:
- 2.71
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: Concentration: 7.81 µM
- Key result
- Run / experiment:
- other: 2
- Parameter:
- other: cell viability [%]
- Value:
- 46.9
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Key result
- Run / experiment:
- other: 2
- Parameter:
- other: EC1.5 [µM]
- Value:
- 2.36
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Other effects / acceptance of results:
- Acceptance Criteria
The test meets acceptance criteria if:
- the luciferase activity induction of the positive control is statistically significant above the threshold of 1.5 (using a t-test) in at least one of the tested concentrations
- the average induction in the three technical replicates for the positive control at a concentration of
64 µM is between 2 and 8
- the EC1.5 value of the positive control is within two standard deviations of the historical mean
- the average coefficient of variation (CV; consisting of 6 wells) of the luminescence reading for the
negative (solvent) control DMSO is <20% in each repetition.
The controls fullfilled the validity criteria of the test. - Conclusions:
- In this study under the given conditions the test item did not induce the luciferase activity in the transgenic KeratinoSens™ cell line in at least two independent experiment runs. Therefore, the test item can be considered as non sensitiser.
The data generated with this method may be not sufficient to conclude on the absence of skin sensitisation potential of chemicals and should be considered in the context of integrated approach such as IATA. - Executive summary:
In the present study dodecyltrimethylammonium bromide was dissolved in DMSO. Based on a molecular weight of 308.34 g/mol a stock solution of 200 mM was prepared.
Based on the stock solution a set of twelve master solutions in 100% solvent was prepared by serial dilution using a constant dilution factor of 1:2. These master solutions were diluted 1:100 in cell culture medium and a stable suspension was formed. The following concentration range was tested in the assay:
2000, 1000, 500, 250, 125, 62.5, 31.25, 15.63, 7.81, 3.91, 1.95, 0.98 µM
Cells were incubated with the test item for 48 h at 37°C. After exposure cells were lysed and luciferase activity was assessed by luminescence measurement.
Microscopically, cytotoxic effects were observed in the whole tested concentration range.
In the first experiment, no significant luciferase induction >1.5 was found in the tested concentration range andno EC1.5 value could be calculated.
In the second experiment, a max luciferase activity (Imax) induction of 2.71 was determined at a test item concentration of 7.81 µM, but the corresponding cell viability was 46.9%. The lowest tested concentration with a significant luciferase induction >1.5 (2.15) was found to be 3.91 µM. The corresponding cell viability was > 70% (72.4%).The calculated EC1.5 was < 1000 µM (2.36 µM).
However, due to the observed cytotoxicity in the whole tested concentration range, it was assumed that the induction above the threshold was caused by cellular stress.A dose response for luciferase activity induction was observed for each individual run as well as for an overall luciferase activity induction. However, this effect was below the threshold value of 1.5.
Under the condition of this study the test item is considered as non sensitiser.
Referenceopen allclose all
Cysteine and Lysine Values of the Calibration Curve
Sample |
Cysteine Peptide |
Lysine Peptide |
||
Peak Area |
Peptide Concentration [mM] |
Peak Area |
Peptide Concentration [mM] |
|
STD1 |
5045.1484 |
0.5340 |
4484.7075 |
0.5340 |
STD2 |
2548.3921 |
0.2670 |
2226.4971 |
0.2670 |
STD3 |
1279.3019 |
0.1335 |
1125.6809 |
0.1335 |
STD4 |
637.0319 |
0.0667 |
549.5435 |
0.0667 |
STD5 |
311.3560 |
0.0334 |
277.2835 |
0.0334 |
STD6 |
153.8533 |
0.0167 |
145.1036 |
0.0167 |
STD7 |
0.0000 |
0.0000 |
0.0000 |
0.0000 |
Depletion of the Cysteine Peptide
Cysteine Peptide |
||||||
Sample |
Peak Area |
Peptide Conc. [mM] |
Peptide Depletion [%] |
Mean Peptide Depletion [%] |
SD of Peptide Depletion [%] |
CV of Peptide Depletion [%] |
Positive Control |
1371.9458 |
0.1446 |
72.07 |
72.27 |
0.27 |
0.38 |
1367.3362 |
0.1441 |
72.17 |
||||
1346.7943 |
0.1419 |
72.58 |
||||
Test Item |
4949.3926 |
0.5227 |
0.00 |
0.10 |
0.11 |
114.56 |
4901.3276 |
0.5176 |
0.22 |
||||
4908.6392 |
0.5184 |
0.07 |
Depletion of the Lysine Peptide
Lysine Peptide |
||||||
Sample |
Peak Area |
Peptide Conc. [mM] |
Peptide Depletion [%] |
Mean Peptide Depletion [%] |
SD of Peptide Depletion [%] |
CV of Peptide Depletion [%] |
Positive Control |
1489.3650 |
0.1777 |
64.50 |
63.51 |
0.86 |
1.35 |
1548.7262 |
0.1848 |
63.08 |
||||
1554.0549 |
0.1854 |
62.95 |
||||
Test Item |
4220.2124 |
0.5031 |
0.00 |
0.00 |
0.00 |
n/a |
4239.8545 |
0.5054 |
0.00 |
||||
4222.2485 |
0.5033 |
0.00 |
n/a: not applicable
Prediction Model 1
Cysteine 1:10/ Lysine 1:50 Prediction Model 1
Mean Cysteine andLysine PPD |
Reactivity Class |
DPRA Prediction² |
0.00% ≤ PPD ≤ 6.38% |
No or Minimal Reactivity |
Negative |
6.38% < PPD ≤ 22.62% |
Low Reactivity |
Positive |
22.62% < PPD ≤ 42.47% |
Moderate Reactivity |
|
42.47% < PPD ≤ 100% |
High Reactivity |
1 The numbers refer to statistically generated threshold values and are not related to the precision of the measurement.
2 DPRA predictions should be considered in the framework of an IATA.
Prediction Model 2
Cysteine 1:10 Prediction Model
Cysteine PPD |
ReactivityClass |
DPRA Prediction² |
0.00% ≤ PPD ≤ 13.89% |
No or Minimal Reactivity |
Negative |
13.89% < PPD ≤ 23.09% |
Low Reactivity |
Positive |
23.09% < PPD ≤ 98.24% |
Moderate Reactivity |
|
98.24% < PPD ≤ 100% |
High Reactivity |
Categorization of the Test Item
Prediction Model |
Prediction Model 1 |
Prediction Model 2 |
||||
Test Substance |
Mean Peptide Depletion [%] |
Reactivity Category |
Prediction |
Mean Peptide Depletion [%] |
Reactivity Category |
Prediction |
Test Item |
0.05 |
Minimal Reactivity |
no sensitiser |
0.10 |
Minimal Reactivity |
no sensitiser |
Positive Control |
67.89 |
High Reactivity |
sensitiser |
72.27 |
Moderate Reactivity |
sensitiser |
Results of the Cytotoxicity Measurement
|
Concentration [µM] |
Cell Viability [%] |
|||
Experiment 1 |
Experiment 2 |
Mean |
SD |
||
Solvent Control |
- |
100 |
100 |
100 |
0.0 |
Positive Control |
4.00 |
102.3 |
102.7 |
102.5 |
0.2 |
8.00 |
101.2 |
104.2 |
102.7 |
2.1 |
|
16.00 |
96.5 |
102.3 |
99.4 |
4.1 |
|
32.00 |
94.2 |
102.3 |
98.3 |
5.7 |
|
64.00 |
81.3 |
105.9 |
93.6 |
17.4 |
|
Test Item |
0.98 |
81.1 |
98.7 |
89.9 |
12.4 |
1.95 |
63.5 |
83.8 |
73.7 |
14.4 |
|
3.91 |
55.6 |
72.4 |
64.0 |
11.9 |
|
7.81 |
35.9 |
46.9 |
41.4 |
7.8 |
|
15.63 |
12.9 |
20.6 |
16.7 |
5.5 |
|
31.25 |
0.2 |
0.5 |
0.4 |
0.2 |
|
62.50 |
0.1 |
0.0 |
0.0 |
0.1 |
|
125.00 |
0.1 |
-0.1 |
0.0 |
0.1 |
|
250.00 |
0.1 |
0.1 |
0.1 |
0.0 |
|
500.00 |
0.3 |
0.3 |
0.3 |
0.0 |
|
1000.00 |
0.3 |
0.3 |
0.3 |
0.0 |
|
2000.00 |
0.4 |
0.9 |
0.6 |
0.3 |
Induction of Luciferase Activity Experiment 1
Experiment 1 |
Concentration [µM] |
Fold Induction |
Significance |
||||
Rep. 1 |
Rep. 2 |
Rep. 3 |
Mean |
SD |
|||
Solvent Control |
- |
1.00 |
1.00 |
1.00 |
1.00 |
0.00 |
|
Positive Control |
4.00 |
1.14 |
1.01 |
1.00 |
1.05 |
0.08 |
|
8.00 |
1.52 |
1.19 |
1.20 |
1.30 |
0.19 |
|
|
16.00 |
1.82 |
1.69 |
1.33 |
1.61 |
0.26 |
* |
|
32.00 |
2.94 |
2.80 |
2.21 |
2.65 |
0.39 |
* |
|
64.00 |
5.54 |
7.93 |
3.94 |
5.80 |
2.01 |
* |
|
Test Item |
0.98 |
0.37 |
0.85 |
0.88 |
0.70 |
0.28 |
|
1.95 |
0.59 |
0.95 |
0.92 |
0.82 |
0.20 |
|
|
3.91 |
0.96 |
1.42 |
1.23 |
1.20 |
0.23 |
|
|
7.81 |
1.06 |
1.25 |
1.11 |
1.14 |
0.10 |
|
|
15.63 |
1.33 |
1.35 |
1.32 |
1.34 |
0.02 |
|
|
31.25 |
0.02 |
0.02 |
0.02 |
0.02 |
0.00 |
|
|
62.50 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
125.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
250.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
500.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
1000.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
2000.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
* = significant induction according to Student’s t-test, p < 0.05
Induction of Luciferase Activity Experiment 2
Experiment 2 |
Concentration [µM] |
Fold Induction |
Significance |
||||
Rep. 1 |
Rep. 2 |
Rep. 3 |
Mean |
SD |
|||
Solvent Control |
- |
1.00 |
1.00 |
1.00 |
1.00 |
0.00 |
|
Positive Control |
4.00 |
1.01 |
0.93 |
1.10 |
1.01 |
0.09 |
|
8.00 |
1.07 |
1.10 |
1.06 |
1.08 |
0.02 |
|
|
16.00 |
1.31 |
1.33 |
1.51 |
1.38 |
0.11 |
|
|
32.00 |
2.02 |
2.22 |
2.02 |
2.08 |
0.12 |
* |
|
64.00 |
3.52 |
3.78 |
3.71 |
3.67 |
0.13 |
* |
|
Test Item |
0.98 |
1.08 |
1.07 |
1.07 |
1.07 |
0.00 |
|
1.95 |
1.24 |
1.38 |
1.35 |
1.33 |
0.07 |
|
|
3.91 |
2.17 |
2.09 |
2.20 |
2.15 |
0.05 |
* |
|
7.81 |
2.67 |
2.67 |
2.79 |
2.71 |
0.07 |
* |
|
15.63 |
2.69 |
2.47 |
2.74 |
2.63 |
0.14 |
* |
|
31.25 |
0.07 |
0.09 |
0.08 |
0.08 |
0.01 |
|
|
62.50 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
125.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
250.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
500.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
1000.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
2000.00 |
0.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
* = significant induction according to Student’s t-test, p < 0.05
Induction of Luciferase Activity – Overall Induction
|
Concentration [µM] |
Fold Induction |
Significance |
|||
Experiment 1 |
Experiment 2 |
Mean |
SD |
|||
Solvent Control |
- |
1.00 |
1.00 |
1.00 |
0.00 |
|
Positive Control |
4.00 |
1.05 |
1.01 |
1.03 |
0.02 |
|
8.00 |
1.30 |
1.08 |
1.19 |
0.16 |
|
|
16.00 |
1.61 |
1.38 |
1.50 |
0.16 |
|
|
32.00 |
2.65 |
2.08 |
2.36 |
0.40 |
* |
|
64.00 |
5.80 |
3.67 |
4.74 |
1.51 |
|
|
Test Item |
0.98 |
0.70 |
1.07 |
0.89 |
0.26 |
|
1.95 |
0.82 |
1.33 |
1.07 |
0.36 |
|
|
3.91 |
1.20 |
2.15 |
1.68 |
0.67 |
|
|
7.81 |
1.14 |
2.71 |
1.93 |
1.11 |
|
|
15.63 |
1.34 |
2.63 |
1.98 |
0.92 |
|
|
31.25 |
0.02 |
0.08 |
0.05 |
0.04 |
|
|
62.50 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
125.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
250.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
500.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
1000.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
|
2000.00 |
0.00 |
0.00 |
0.00 |
0.00 |
|
* = significant induction according to Student’s t-test, p < 0.05
Additional Parameters
Parameter |
Experiment 1 |
Experiment 2 |
Mean |
SD |
EC1.5[µM] |
n.a. |
2.36 |
n.a. |
n.a. |
Imax |
1.34 |
2.71 |
2.02 |
0.97 |
IC30[µM] |
1.59 |
4.28 |
2.93 |
1.90 |
IC50[µM] |
5.01 |
7.34 |
6.17 |
1.65 |
n.a.:not applicable
Acceptance Criteria
Criterion |
Range |
Experiment 1 |
pass/fail |
Experiment 2 |
pass/fail |
CV Solvent Control |
< 20% |
14.0 |
pass |
9.0 |
pass |
No. of positive control concentration steps with significant luciferase activity induction >1.5 |
≥ 1 |
3.0 |
pass |
2.0 |
pass |
EC1.5 PC |
7 < x < 34 µM |
13.11 |
pass |
18.67 |
pass |
Induction PC at 64 µM |
2.00 < x < 8.00 |
5.80 |
pass |
3.67 |
pass |
Historical Data
Acceptance Criterion |
Range |
Mean |
SD |
N |
CV Solvent Control |
< 20% |
11.3 |
3.3 |
41 |
No. of positive control concentration steps with significant luciferase activity induction >1.5 |
≥ 1 |
2.3 |
0.6 |
41 |
EC1.5 PC |
7 < x < 34 µM |
20.4 |
6.7 |
41 |
Induction PC at 64 µM |
2.00 < x < 8.00 |
3.3 |
1.1 |
41 |
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (not sensitising)
- Additional information:
For the evaluation of the skin sensitisation potential of the test substance a Weight of Evidence approach was used. An in silico assessment using Derek Nexus v5.0 was carried out. The protein reactivity of the test item was assessed in a Direct Peptide Reactivity Assay (DPRA) and a Keratinocyte Activation Assay (KeratinoSens) was performed to evaluate the keratinocyte activation potential.
In silico assessment (reference 7.4.1-1)
For in silico assessment, Derek Nexus v5.0 was used. Skin sensitising properties were regarded as equivocal based on the described QSAR method (Derek, 2018).
DPRA (reference 7.4.1-2)
In the present study dodecyltrimethylammonium bromide was dissolved in acetonitrile, based on the results of the pre-experiments. Based on a molecular weight of 308.34 g/mol a 100 mM stock solution was prepared. The test item solutions were tested by incubating the samples with the peptides containing either cysteine or lysine for 24 ± 2 h at 25 ± 2.5 °C. Subsequently samples were analysed by HPLC.
For the 100 mM solution of the test item no turbidity or precipitation was observed when diluted with the cysteine peptide solution. After the 24 h±2 h incubation period but prior to the HPLC analysis samples were inspected for precipitation, turbidity or phase separation. No precipitation, turbidity or phase separation was observed for any of the samples.
For the 100 mM solution of the test item no turbidity or precipitation was observed when diluted with the lysine peptide solution. After the 24 h±2 h incubation period but prior to the HPLC analysis samples were inspected for precipitation, turbidity or phase separation. Phase separation was observed for the co-elution control of the positive control. No precipitation, turbidity or phase separation was observed for the samples of the test item. Samples were not centrifuged prior to the HPLC analysis.
Since the acceptance criteria for the depletion range of the positive control were fulfilled, the observed phase separation was regarded as insignificant.
No co-elution of test item with the peptide peaks was observed. Sensitising potential of the test item was predicted from the mean peptide depletion of both analysed peptides (cysteine and lysine) by comparing the peptide concentration of the test item treated samples to the corresponding reference control C (RC C acetonitrile).
The 100 mM stock solution of the test item showed minimal reactivity towards the synthetic peptides. The mean depletion of both peptides was ≤ 6.38% (0.05%). Based on the prediction model 1 the test item can be considered as non-sensitiser.
The 100 mM stock solution of the positive control (cinnamic aldehyde) showed high reactivity towards the synthetic peptides. The mean depletion of both peptides was 67.89%.
The controls confirmed the validity of the study for both the cysteine and lysine run.
KeratinoSens (reference 7.4.1-3)
In the present study dodecyltrimethylammonium bromide was dissolved in DMSO. Based on a molecular weight of 308.34 g/mol a stock solution of 200 mM was prepared.
Based on the stock solution a set of twelve master solutions in 100% solvent was prepared by serial dilution using a constant dilution factor of 1:2. These master solutions were diluted 1:100 in cell culture medium and a stable suspension was formed. The following concentration range was tested in the assay:
2000, 1000, 500, 250, 125, 62.5, 31.25, 15.63, 7.81, 3.91, 1.95, 0.98 µM
Cells were incubated with the test item for 48 h at 37°C. After exposure cells were lysed and luciferase activity was assessed by luminescence measurement.
Microscopically, cytotoxic effects were observed in the whole tested concentration range.
In the first experiment, no significant luciferase induction >1.5 was found in the tested concentration range and no EC1.5 value could be calculated.
In the second experiment, a max luciferase activity (Imax) induction of 2.71 was determined at a test item concentration of 7.81 µM, but the corresponding cell viability was 46.9%. The lowest tested concentration with a significant luciferase induction >1.5 (2.15) was found to be 3.91 µM. The corresponding cell viability was > 70% (72.4%).The calculated EC1.5 was < 1000 µM (2.36 µM).
However, due to the observed cytotoxicity in the whole tested concentration range, it was assumed that the induction above the threshold was caused by cellular stress.A dose response for luciferase activity induction was observed for each individual run as well as for an overall luciferase activity induction. However, this effect was below the threshold value of 1.5.
Under the condition of this study the test item is considered as non sensitiser.
Conclusion
Skin sensitising properties were regarded as equivocal based on the described QSAR method (Derek, 2018). Based on the results of an in chemico/in vitro test strategy the test item is not peptide reactive (DPRA,OECD TG 442C,reference 7.4.1-2) and does not activate keratinocytes (Keratinosens, OECD TG 442D, reference 7.4.1-1). Therefore, the substance is not predicted to be a skin sensitizer. The overall result is based on two concordant findings. Considering an AOP “2 out of 3” the test item is predicted as non-skin sensitizer and further tests are not necessary.
Respiratory sensitisation
Endpoint conclusion
- Endpoint conclusion:
- no study available
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. Based on available data on skin sensitisation, the test item is not classified as a skin sensitizer according to Regulation (EC) No 1272/2008 (CLP), as amended for the tenth time in Regulation (EU) No 2017/776.
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