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Description of key information

The key information is based on three different in-vitro/in-silico tests addressing different events of the adverse outcome pathway (AOP), namely activation of keratinocytes (KeratinoSens™ assay), the molecular initiating event being protein reactivity (in-chemico direct peptide reactivity assay), and dendritic cell activation (h-CLAT).

Key value for chemical safety assessment

Skin sensitisation

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

In the in-vitro KeratinoSens™ assay, the following concentration range was tested: 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. In two separate experiments, no significant luciferase induction > 1.5 was found in the tested concentration range. Therefore, no EC1.5 value could be calculated. No dose response for luciferase activity induction was observed for each individual run as well as for an overall luciferase activity induction. Under the condition of this study the test item is therefore considered as non sensitiser.

In the in-chemico direct peptide reactivity assay (DPRA), 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. 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 test item samples. 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. 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% (1.06%). 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 with a mean depletion of both peptides of 65.83%.

In the human cell line activation test (h-CLAT), 2-(n-ethylanilino)ethanol was dissolved in DMSO.For the dose finding assay stock solutions with concentrationsranging from 500 mg/mL to 3.91 mg/mLwere prepared by a serial dilution of 1:2. Cells were incubated with the test item for 24 h at 37°C. After exposure cells were stained with propidium iodide and cell viability (CV) was measured by FACS analysis. A CV75 of 578.15 ± 15.95 µg/mL was derived in the dose finding assay. Based on the CV75, the main experiment was performed covering the following concentration steps: 693.77, 578.15, 481.79, 401.49, 334.57, 278.81, 232.34, 193.62 µg/mL In all experiments no precipitation or turbidity of the test item was observed for all concentration steps when mixing the test item stock solutions with cell culture medium. Cells were incubated with the test item for 24 h at 37°C. After exposure cells were stained and cell surface markers CD54 and CD86 were measured by FACS analysis. Cell viability was assessed in parallel using propidium iodide staining. Cytotoxic effects were observed for the cells treated with the test item. Relative cell viability at the highest test item concentration was reduced to 60.2% (CD86), 57.2% (CD54) and 56.7% (isotype IgG1 control) in the first experiment and to 55.9% (CD86), 56.4% (CD54) and 55.6% (isotype IgG1 control) in the second experiment. In the first experiment the expression of the cell surface markers CD86 and CD54 was not upregulated above the threshold of 150% and 200%. In the second experiment the expression of the cell surface marker CD86 was upregulated to 181% at a concentration of 693.77 µg/mL. This was the only upregulation above the threshold of 150%, no other upregulation was observed. Since there was no upregulation in the first and the one upregulation in the second experiment, the outcome of the study must be considered as inconclusive.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Based on the three guideline tests performed, 2-(n-ethylanilino)ethanol did not appear to be a skin sensitizer.

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