1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-coco acyl derivs., hydroxides, inner salts

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

02 May, 2018 - 04 May, 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

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

An assessment found the test item was able to directly reduce MTT and an additional procedure using freeze-killed tissues was performed. However, the results obtained showed that no interference due to direct reduction of MTT occurred.

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-coco acyl derivs., hydroxides, inner salts

EC Number:

268-761-3

EC Name:

1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-coco acyl derivs., hydroxides, inner salts

Cas Number:

68139-30-0

Molecular formula:

C9H19N2O5S(CnH(2n+1)), (n=7,9,11,13,15,17)

IUPAC Name:

1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-coco acyl derivs., hydroxides, inner salts

Test material form:

solid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: DSP-001.
- Expiration date of the lot/batch: 15 August 2018.
- Purity test date: No data.

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature in the dark.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The test item (solid) was pressed flat and appropriate sized discs were cut out. The application of the discs is an acceptable method in accordance with the MatTek SOP MK-24-007-0024 of the validated and regulatory accepted EpiDerm SCT.

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

foreskin from a single donor

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ Reconstructed Human Epidermis Model Kit.
- Supplier: MatTek.
- Tissue batch number(s): 28602.
- Delivery date: 01 May 2018.
- Date of initiation of testing: 02 May 2018.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37 °C.
- Temperature of post-treatment incubation (if applicable): Room temperature.

REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps:
- Observable damage in the tissue due to washing:
- Modifications to validated SOP:

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1.0 mg/mL.
- Incubation time: 3 hours.
- Spectrophotometer: Labtech LT-4500 microplate reader.
- Wavelength: 570 nm.
- Linear OD range of spectrophotometer: 0-4 OD.

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: ET-50 fall within the range of the 1996 EpiDerm database of 4.77 – 8.72 hours.
- Barrier function: Lipid precursors used to enhance epidermal barrier formation.
- Morphology: Good retention of epidermal morphology for up to 3 weeks.
- Contamination: All tissues are visually inspected and if physical imperfections are noted, tissues are rejected for shipment.
- Reproducibility: Tissues may be stored at 4°C for up to 6 days prior to use, although best reproducibility obtained if used on same day.

NUMBER OF REPLICATE TISSUES: 2.

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Fresh tissues / killed tissues : The test item was shown to directly reduce MTT in the direct MTT reduction test, therefore the determination of skin corrosion potential was performed in parallel on viable and freeze-killed tissues.
- Procedure used to prepare the killed tissues: Freeze-killed tissues were prepared prior to the study by placing untreated EPIDERM tissues in an empty 12-well plate and storing in a freezer (−14 to −30 °C) for a minimum of 24 hours. Before use each tissue was thawed by placing in 0.9 mL of assay medium for approximately 1 hour at room temperature.
- Number of replicates: 2.
- Method of calculation used: Relative mean viability (%) - (mean OD570 of test item / mean OD570 of negative control) x 100.

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

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: 0.0931 g.

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): 50 µL.
- Concentration (if solution): 100%.

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL.
- Concentration (if solution): 100%.

Duration of treatment / exposure:

3 and 60 minutes.

Number of replicates:

2

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minute exposure

Value:

90

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

Percentage viability 100%

Positive controls validity:

valid

Remarks:

Percentage viability 3.3%

Remarks on result:

no indication of irritation

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minute exposure

Value:

80.8

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

Percentage viability 100%

Positive controls validity:

valid

Remarks:

Percentage viability 2.9%

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: None observed.
- Direct-MTT reduction: The results obtained from the freeze-killed tissues showed that no interference due to direct reduction of MTT occurred. It was therefore considered unnecessary to use the results for quantitative correction of results or for reporting purposes.
- Colour interference with MTT: The solution containing the test item did not become colored. This was taken to indicate the test item did not have the potential to cause color interference.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The mean OD570 for the negative control treated tissues was 1.854 for the 3-Minute exposure period and 1.927 for the 60-Minute exposure period. The negative control acceptance criteria were therefore satisfied.
- Acceptance criteria met for positive control: The relative mean tissue viability for the positive control treated tissues was 2.9% relative to the negative control following the 60-Minute exposure period. The positive control acceptance criterion was therefore satisfied.
- Acceptance criteria met for variability between replicate measurements: In the range 20 to 100% viability the Coefficient of Variation between the two tissue replicates of each treatment group did not exceed 30%. The acceptance criterion was therefore satisfied.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The test item was considered to be non-corrosive to the skin.

Executive summary:

1-Propanaminium, N-(3-aminopropyl)-2-hydroxy- N,N-dimethyl-3-sulfo-, N-coco acyl derivs., hydroxides, inner salts was tested according to OECD Test Guideline 431 to predict its potential for skin corrosivity by cytotoxicity in the EpiDerm™ tissue. Following a 60 minute exposure period to the test item, mean tissue viability was 80.8%, well above the <15% threshold for classification as corrosive to the skin.