Condensation products of N-(6-aminohexyl)-12-hydroxyoctadecanamide and sebacic acid

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07 to 23 March 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Condensation products of N-(6-aminohexyl)-12-hydroxyoctadecanamide and sebacic acid

EC Number:

945-990-3

IUPAC Name:

Condensation products of N-(6-aminohexyl)-12-hydroxyoctadecanamide and sebacic acid

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source of test material: Kyoeisha Chemical Co., Ltd
- Lot No.of test material: 6033175
- Expiration date of the lot/batch: 17 March 2019
- Purity test date: 3 October 2017

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature (Ambient), in original container as supplied by the Sponsor. Kept in tightly closed container and away from heat or sunlight
- Stability under test conditions: Assumed stable for the duration of the study
- Solubility and stability of the test substance in the solvent/vehicle: Test item administed as supplied, no stability of test item in the vehicle required.
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: None

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Test substance administered as supplied.

In vitro test system

Test system:

human skin model

Remarks:

SkinEthicTM RHE model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: human derived non-transformed epidermal keratinocytes

Justification for test system used:

This study addresses the human health endpoint skin corrosion. It makes use of reconstructed human epidermis (RHE) (human derived non-transformed epidermal keratinocytes) which closely mimics the histological, morphological, biochemical and physiological properties of the upper parts of the human skin. The use of reconstructed human epidermis (RHE) is also recommended by the OECD and other regulatory authorities. The SkinEthicTM RHE model has been validated and is part of the OECD validated reference methods (VRMs), it is a recommended model for conducting in vitro skin corrosion studies. The results of the study are believed to be of value in predicting the potential of inducing skin corrosivity by the test item in humans.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: SkinEthicTM RHE model
- Tissue batch number: 18-RHE-029
- Production date: Not specified
- Shipping date: Not specified
- Delivery date: Not specified
- Date of initiation of testing: 10 March 2018

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 3 minutes at room temperature and 60 minutes at 37±1 °C in 5±1% CO2 in 95% humidified atmosphere
- Temperature of post-treatment incubation (if applicable): Not applicable

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: After exposure, tissues were rinsed and then dried with cotton buds. Any test item was initially removed by knocking the treated insert on the beaker or by holding upside down with forceps. Treated tissues were rinsed 20 times in a constant soft stream of 1 mL DPBS at a distance of
5-8 cm from the insert to remove all residual test item from the epidermal surface. Nylon mesh (applied on the negative control and positive control) was removed by washing. The bottom of tissue inserts were dried on a sterile absorbent paper (Kim wipes) for 1-2 seconds. The surface of the stratum corneum was gently swept with both ends of a cotton tip (5-6 turns per end). After washing, inserts were transferred to a holding plate containing 300 µL maintenance medium.
- Observable damage in the tissue due to washing: None
- Modifications to validated SOP: None

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: Incubated for 180 minutes at 37±1 °C in 5±1% CO2 in a 95% humidified atmosphere
- Spectrophotometer: SynergyHT Microplate Reader
- Wavelength: 570 nm.
- Filter: not specified
- Filter bandwidth: 570±30 nm
- Linear OD range of spectrophotometer: not specified

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: In the range of 20-100% viability and for ODs ≥ 0.3, difference of viability between the tissue replicates was not > 30%.
- Barrier function: yes
- Morphology: mimics the histological, morphological, biochemical and physiological properties of the upper parts of the human skin
- Contamination: None
- Reproducibility: yes, within acceptable parameters

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Fresh tissues / killed tissues: The test item did not produce direct MTT reduction when compared to the concurrent negative control (maintenance medium).

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: One main test

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be corrosive to skin if the viability after 3 minutes exposure is less than 50%, or if the viability after 3 minutes exposure is greater than or equal to 50% and the viability after 1 hour exposure is less than 15%.
- The test substance is considered to be non-corrosive to skin if the viability after 3 minutes exposure is greater than or equal to 50% and the viability after 1 hour exposure is greater than or equal to 15%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): , the epidermal surface of tissue was wetted by addition of 20 µL sterile distilled water using a positive displacement micropipette followed by application of 20 mg ± 3 mg of test item/0.5 cm2
- Concentration (if solution):

VEHICLE
- Amount(s) applied (volume or weight with unit):
- Concentration (if solution):
- Lot/batch no. (if required):
- Purity:

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 40 µL/0.5 cm2 of sterile distilled water was applied
- Concentration (if solution):

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 40 µL/0.5 cm2 of 8N KOH was applied
- Concentration (if solution):

Duration of treatment / exposure:

For 3 minutes at room temperature and 60 minutes at 37±1 °C in 5±1% CO2 in 95% humidified atmosphere

Number of replicates:

3

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minutes exposure (Negative control)

Value:

100

Negative controls validity:

valid

Remarks:

Sterile distilled water

Remarks on result:

no indication of irritation

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minutes exposure (Negative control)

Value:

100

Negative controls validity:

valid

Remarks:

Sterile distilled water

Remarks on result:

no indication of irritation

Results 3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minute exposure (Test item)

Value:

90.7

Remarks on result:

no indication of irritation

Results 4

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minute exposure (Test item)

Value:

65.4

Remarks on result:

no indication of irritation

Results 5

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minutes exposure (Positve control)

Value:

0.1

Positive controls validity:

valid

Remarks:

8N KOH

Remarks on result:

positive indication of irritation

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: None
- Direct-MTT reduction: Test item did not produce direct MTT reduction when compared to the concurrent negative control (maintenance medium)
- Colour interference with MTT: The results of the color interference test shows no interference in optical density due to the test item

DEMONSTRATION OF TECHNICAL PROFICIENCY: All criteria for a valid study were met.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes
- Acceptance criteria met for positive control: yes
- Acceptance criteria met for variability between replicate measurements: yes
:

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

From the results of this study, it is concluded that N, N'-bis {6-[12-(hydroxy) octadecanoylamino] hexyl} decanediamide is non-corrosive in accordance with the United Nations Globally Harmonized System of Classification and Labelling of Chemicals as indicated in OECD Test Guideline 431 under the specified conditions of this study.