Reaction products of stearic acid, diethylenetriamine and N-aminoethylethanolamine and acetic acid

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

08/02/2018 - 03/04/2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 442D (In Vitro Skin Sensitisation: ARE-Nrf2 Luciferase Test Method)

Deviations:

no

Remarks:

see principles of method below for full details

Principles of method if other than guideline:

Deviations from the guidelines:
- Standard states MTT musted be used for an incubation period of 4 hours, solubilised overnight in 10% SDS and read at 690 nm, the study uses MTT for a 3 hr incubation period, which is solubilised in isopropanol and read at 570 nm.
- The standard states that cinnamic aldehyde must be used at 4 - 64 μM, whereas the study uses cinnamic aldehyde 8 - 128 μM.

GLP compliance:

yes (incl. QA statement)

Type of study:

activation of keratinocytes

Justification for non-LLNA method:

The test (KeratinoSens) test is a method for which validation studies have been completed followed by an independent peer review conducted by the European Union Reference Laboratory for Alternatives to Animal Testing (EURL ECVAM) and is considered scientifically valid when used as part of an integrated approach to testing to determine skin sensitisers and non-sensitisers for the purposes of hazard classification and labelling.

Test material

Test material information

Constituent 1

Reference substance name:

Reaction products of stearic acid, diethylenetriamine and N-aminoethylethanolamine and acetic acid

IUPAC Name:

Reaction products of stearic acid, diethylenetriamine and N-aminoethylethanolamine and acetic acid

Test material form:

solid: bulk

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 0001230562
- Expiration date of the lot/batch: 08/06/2019
- Purity test date: N/A

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Assumed to be stable
- Stability under test conditions:Assumed to be stable
- Solubility and stability of the test substance in the solvent/vehicle: Assumed to be stable
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: Assumed to be stable

In vitro test system

Details on the study design:

Skin sensitisation (In vitro test system) - Details on study design: Using a test system (KeratinoSens cell line (test system) derived from the HaCaT human keratinocytes) a luciferase reporter gene transcriptionally controlled by the Anti-oxidant Response Element from a gene that is known to be up regulated in contact with sensitisers was quantitatively measured. Experiments were performed in plates where cells were exposed to in individual concentrations of the test substance or appropriate control substances. Test concentrations were determined based on the solubility of the test item in ethanol (max. 20 mg/mL in Ethanol), and for subsequent dilution in cell culture medium a top concentration of 200 μg/mL) with 3 replicates per concentration per treatment.

Results and discussion

Positive control results:

The positive control, 'Cinnamic Aldehyde' showed that there was a dose-dependent increase of induction, which was > 1.5 - fold in at least one concentration and the CV% of blank values was < 20% so was accepted. The positive control was positive at both 64 and 129 μM.

In vitro / in chemico

Results

Results 1

Key result

Run / experiment:

other: Test Item

Parameter:

other: EC1.5

Value:

1.5

Vehicle controls validity:

not valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of skin sensitisation

Results 2

Run / experiment:

other: Test item

Parameter:

other: Imax

Value:

0.854

Vehicle controls validity:

not specified

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of skin sensitisation

Other effects / acceptance of results:

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Not specified.
- Acceptance criteria met for positive control: Not all criteria met, however considered valid based on Positive Control results.
- Acceptance criteria met for variability between replicate measurements: No specified.
- Range of historical values if different from the ones specified in the test guideline: Same.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The Reaction products of stearic acid, diethylenetriamine and N-aminoethylethanolamine and acetic acid was classified as a Non-sensitiser to human skin as EC1.5 value value did not exceed the minimum threshold (1.5) and the Imax value was between 0.854 - 1.183.

Executive summary:

The study assesses the In vitro sensitisation potential of Reaction products of stearic acid, diethylenetriamine and N-aminoethylethanolamine and acetic acid using the test system (KeratinSens test method). The study is GLP compliant and is performed according to the OECD 442D guidelines. The experiment use the quantification of luciferase expression in the cell reporter line to determine the response to the test item, a negative control and a positive control, reporting the EC1.5 and Imax values. In all test item experiments the EC1.5 value did not exceed the minimum threshold value (1.5) and the Imax value range was 0.854 - 1.183, which lead to conclusion that the test item is not a skin sensitiser (GHS criteria determined).