Disodium Capryloyl Glutamate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 17 May 2022 to 20 May 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2022

Report date:

2022

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Version / remarks:

14 June 2021

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Reaction mass of L-Glutamic acid, N-(1-oxooctyl)-, sodium salt and N-L-glutamyl-L-glutamic acid, N'-(1-oxooctyl)-, sodium salt

Cas Number:

167888-81-5

Molecular formula:

Not applicable, see single constituents

IUPAC Name:

Reaction mass of L-Glutamic acid, N-(1-oxooctyl)-, sodium salt and N-L-glutamyl-L-glutamic acid, N'-(1-oxooctyl)-, sodium salt

Test material form:

liquid

Details on test material:

The product identified with the Registration Number 01-2120781106-56-0000 is always been manufactured in the same conditions with same raw materials (quality and quantity) and synthesis reaction.
So all tests done before ECHA discussion about name and identificative are still with the old name but they refer in any case to the registered molecule. The new name was changed after ECHA contact in order to find the most proper identificative for the molecule.

Specific details on test material used for the study:

Batch no.: 2044 supplied by the sponsor
Aspect: clear to light turbid liquid
Purity: 33.2%
pH as it is 7.1
Water solution
Storage condition of test material: room temperature (15 °C - 25 °C), stable for at least 12 months
Stability and homogeneity of the test material in the vehicle/solvent under test conditions (e.g. in the exposure medium): stable for 12 months minimum
Solubility: soluble and stable in water
Reactivity of the test material with the incubation material used (e.g. plastic ware): not reactive
More information can be found on the attached report

In vitro test system

Test system:

artificial membrane barrier model

Cell type:

non-transformed keratinocytes

Cell source:

foreskin from multiple donors

Vehicle:

water

Remarks:

Substance in already in water solution

Details on test system:

The test item was preliminary tested to evaluate its interference with MTT endopoint. 16µL of the test item were put in contact with 0.3 ml of MTT and incubated for 180 ± 5 minutes at 37°C and 5% CO2. A negative control (16 µ1 of sterile deionized water) was run concurrently. MTT solution color did not change its color into blue/purple, indicating that the test item was not reducing the MTT.

Furthermore the test item was preliminary tested to evaluate its colorant properties.

10µ,L of the test item were put in contact with 0.09m1 of water and incubated for 30 minutes at room temperature after shaking. A negative control (10 µl of sterile deionized water) was run concurrently. The absorbance was measured with a plate reader equipped with the MTT measurement wavelength filter. After subtraction of the OD of negative control, the OD of the test item solution was < 0.1 (-0.0018, See Annex 05 — Raw data test item colorant properties), indicating that the test item does not interact with the MTT measurement.

In the definitive test, after over-night incubation in the growth medium, the epidermis units were treated with the test item. 16µL of test item, and 16µ1 of positive (SDS 5%) and negative (PBS without Ca and Mg) controls were applied on epidermis units in three replicates. The exposure was carried out for 42 minutes at room temperature. At the end of the exposure period sample and controls were removed with multiple washings with PBS and the tissues were further incubated in 2m1 of growth medium at 37°C, 5% CO2 for 42 h. At the end of the incubation, the viability assay was performed to evaluate the cell survival in the epidermis units. Epidermis units were treated with 0,3mL 1 mg/ml MTT solution (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) for 3 h at 37°C.
The solution was then removed and replaced with isopropanol, with further 2 h incubation at room temperature under medium speed shaking.

2 aliquots of isopropanol were sampled for each epidermis unit and transferred to a 96 well plate for the reading (6 readings total for the test item, positive and negative control).

The absorbance was read with a microplate spectrophotometer equipped with the 570 nm filter. The absorbance values were corrected by subtracting the reading of the blanks (diluent only).

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

16 microliter of test item was applied in three replicates

Duration of treatment / exposure:

42 minutes at room temperature

Number of replicates:

3

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3

Value:

116.15

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

2

Value:

103.36

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Results 3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1

Value:

104.55

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

Acceptance criteria

For the blank (isopropanol): OD has to be < 0.1 for each replicates
For negative control (NC): the mean OD value of the 3 tissues has to be ≥ 0.8 and ≤ 3.0

the standard deviation of the viability mean percentage has to be ≤ 18%



For positive control (CP): the viability mean 8expresses as % versus the NC) has to be < 40%

the standard deviation has to be ≤ 18%



For the test item: the standard deviation of the viability mean percentage has to be ≤ 18%

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The test item Reaction mass of L-Glutamic acid, N-(1-oxooctyl)-, sodium salt and N-L-glutamyl-L-glutamic acid, N'-(1-oxoocty1)-, sodium salt Batch 2044, ABICH code S012­22 tested on Reconstructed Human Epidermis according to OECD 439 and in compliance to GLP, results to be not irritant to the skin (NO CATEGORY), because its mean cell viability is higher than 50%. All acceptance criteria were passed.

In conclusion, it can be stated that, in the experimental conditions here described and according to this result, the test item Reaction mass of L-Glutamic acid, N-(1-oxooctyl)-, sodium salt and N-L-glutamyl-L-glutamic acid, N'-(1-oxoocty1)-, sodium salt Batch 2044, ABICH code S012-22 can be classified as NOT IRRITANT (NO CATEGORY according to UN GHS CLASSIFICATION).

Executive summary:

Purpose of the study is to evaluate the skin irritation potential on Reconstructed Human Epidermis (RHE SkinEthic) of the test item "Reaction mass of L-Glutamic acid, N-(1-oxoocty1)-, sodium salt and N-L-glutamyl-L-glutamic acid, N'-(1-oxoocty1)-, sodium salt", ABICH code S012-22 according to the method described in OECD 439 and in compliance with Good Laboratory Practice (GLP).

This method uses human artificial skin models to assess the skin irritation of chemical substances and mixtures and to properly label them to this respect, if applicable.

The test is based on the evaluation of cell survival after the exposure to the test item through MTT assay and by comparison with epidermis treated with phosphate buffer only (negative control). The MTT method is a colorimetric assay that allows to determinate the percentage of cells alive within an in vitro cultured tissue. This assay is based on the ability of the mitochondrial succinate dehydrogenase enzyme to metabolise the nitro-blue tetrazolium salt, giving a coloured compound that can be measured by spectrophotometer reading.

The test item "Reaction mass of L-Glutamic acid, N-(1-oxooctyl)-, sodium salt and N-L-glutamyl-L-glutamic acid, N'-(1-oxoocty1)-, sodium salt", ABICH code S012-22, in the experimental conditions here described, showed a mean cell survival of 108.02% (±7.07), so it resulted a NOT SKIN IRRITANT (UN-GHS — no Category), because its cell viability is higher than 50% .

The positive control Sodium Doceyl Sulphate (SDS) showed a mean cell viability of 1.58% (±0.03), so it gave the expected results. All the acceptance criteria were passed.