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Description of key information

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.

Based on the results of this study, the classification for N, N’-bis {6-[12-(hydroxy) octadecanoylamino] hexyl} decanediamide is as follows:

Globally Harmonized System of Classification and Labelling of Chemicals : No Category (Non Skin Irritant)

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Referenceopen allclose all

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
Qualifier:
according to
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)
Deviations:
no
GLP compliance:
yes (incl. certificate)
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.
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
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
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
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
3 minute exposure (Test item)
Value:
90.7
Remarks on result:
no indication of irritation
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
60 minute exposure (Test item)
Value:
65.4
Remarks on result:
no indication of irritation
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
:
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.
Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10 to 23 March 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Deviations:
not specified
Qualifier:
according to
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Deviations:
not specified
GLP compliance:
yes (incl. certificate)
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. Container was kept tightly closed 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 iitem administered 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: None, test item administered as supplied


Test system:
human skin model
Remarks:
SkinEthicTM RHE model
Source species:
human
Cell type:
non-transformed keratinocytes
Justification for test system used:
This study addresses the human health endpoint skin irritation. It uses reconstructed human epidermis (RhE) (obtained from 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 recommended by the OECD and other regulatory authorities. SkinEthicTM RHE model has been validated and is part of the OECD validated reference methods (VRMs) and is a recommended model for conducting in vitro skin irritation studies. The results of the study are believed to be of value in predicting the potential of inducing skin irritation by the test item in humans.
Vehicle:
unchanged (no vehicle)
Details on test system:
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: The 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: incubated for 42 minutes at room temperature.
- Temperature of post-treatment incubation (if applicable): After washing and drying, tissues were incubated in a 6-well plate containing 2 mL growth medium at 37 ± 1°C in 5 ± 1% CO2 in a humidified incubator for 42 hours.

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: After exposure, tissues were rinsed and then dried with cotton buds. The test item was removed by rinsing 25 times in a constant soft stream of 1 mL DPBS at 5-8 cm distance from the insert to remove test item from the epidermal surface. Mesh (applied on negative and positive control tissues) 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 up using both ends of a cotton tip (5-6 turns per end). After washing, inserts were transferred to holding plates 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: 180 minutes at 37 ± 1°C in 5 ± 1% CO2 in a 95% humidified incubator.
- Spectrophotometer: SynergyHT Microplate Reader
- Wavelength: 570 nm ± 30 nm
- Filter: not specified
- Filter bandwidth: 570 nm ± 30 nm.
- Linear OD range of spectrophotometer: Not specified

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: Observed results are within the acceptance/historical range of SkinEthic Laboratories/OECD TG acceptance criteria.
- 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
Test item did not produce direct MTT reduction when compared to the concurrent negative control (maintenance media).

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: one main study

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be irritant to skin if the viability after 42 minutes exposure and post-treatment incubation is less than or equal (≤) to 50%.
- The test substance is considered to be non-corrosive to skin if after 42 minutes exposure and post-treatment incubation is more than 50%
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
- Amount(s) applied (weight with unit): 16 ± 2 mg of test item/0.5 cm2

NEGATIVE CONTROL
- Amount(s) applied (volume): 16 µL/0.5 cm2 sterile dulbecco’s phosphate buffered saline (DPBS)

POSITIVE CONTROL
- Amount(s) applied (volume): 16 µL/0.5 cm2 of 5% sodium dodecyl sulphate (5% aq.)
Duration of treatment / exposure:
42 minutes
Duration of post-treatment incubation (if applicable):
42 hours
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
42 minute exposure (Negative control)
Value:
100
Negative controls validity:
valid
Remarks:
(Dulbecco’s Phosphate Buffered Saline (DPBS))
Remarks on result:
no indication of irritation
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
42 minute exposure (Test item)
Value:
101.6
Remarks on result:
no indication of irritation
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
42 minute exposure (Positive control)
Value:
1.5
Positive controls validity:
valid
Remarks:
Sodium dodecyl sulphate (5% aq.)
Remarks on result:
positive indication of irritation
Other effects / acceptance of results:
- OTHER EFFECTS:
- Visible damage on test system: None
- Direct-MTT reduction:
- Colour interference with MTT:

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
Interpretation of results:
GHS criteria not met
Conclusions:
Based on the results of this study, the classification for N, N’-bis {6-[12-(hydroxy) octadecanoylamino] hexyl} decanediamide is as follows:
Globally Harmonized System of Classification and Labelling of Chemicals : No Category (Non Skin Irritant)

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Jan to July 2018
Reliability:
1 (reliable without restriction)
Qualifier:
according to
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Vehicle:
other: corn oil
Controls:
yes, concurrent vehicle
yes, concurrent positive control
yes, concurrent negative control
Amount / concentration applied:
750 μL of the prepared test item [20% (w/v) concentration in corn oil],positive control [750 μL of 20% (w/v) concentration in normal saline], two set of corneas served as negative control and was treated with 750 μL of normal saline and corn oil,
Duration of treatment / exposure:
4 h ± 5 minutes at 32 ± 1 °C
Duration of post- treatment incubation (in vitro):
4 h ± 5 minutes at 32 ± 1 °C
Number of animals or in vitro replicates:
3 corneas per test group. 12 in total
Irritation parameter:
cornea opacity score
Value:
> -0.05 - < 0.19
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Interpretation of results:
GHS criteria not met
Conclusions:
The substance is not irritatiing.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for classification or non-classification

Studies show that this substance is not classified for skin irritation/corrosion or eye irritation.