Registration Dossier

Administrative data

Endpoint:
skin irritation / corrosion, other
Remarks:
Skin corrosivity and irritation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2018-01-10 to 2018-03-26
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)
Version / remarks:
adopted 29 July 2016
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Version / remarks:
adopted 28 July 2015
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
- CAS number: 39464-70-5
- Appearance: Clear, yellow-amber liquid
- Purity: 100%
- Storage conditions: Room temperature (15-25 °C, ≤ 70 % relative humidity (RH))
- Safety precautions: Routine safety precautions (gloves, goggles, face mask, lab coat) for unknown materials were applied to assure personnel health and safety.

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Justification for test system used:
The corrosivity and irritation potential of a chemical may be predicted by measurement of its cytotoxic effect, as reflected in the MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, Thiazolyl blue; CAS number 298-93-1] assay, on EPISKIN^TM(SM) reconstituted human epidermis. This method is approved by international regulatory agencies as a replacement for the identification of irritants / corrosives in the in vivo Rabbit skin assay (OECD No. 404) and is specifically approved as a replacement for the in vivo skin corrosivity and irritation tests within OECD No. 431. and OECD No. 439.
Vehicle:
unchanged (no vehicle)
Details on test system:
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EPISKIN (SM)
- Tissue batch number(s): 18-EKIN-002
- Expiry date: 15 January 2018

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 0.3 mg / mL
- Incubation time: 3 h + / - 5 min
- Wavelength: 570 nm

NUMBER OF REPLICATE TISSUES: two for the corrosion test and three for the irritation test

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- N. of replicates : Two negative controls and two positive controls were run in corrosivity testing and three negative controls and three positive controls were run in irritation testing. As the test item was coloured, two additional test item-treated living tissues were used for the non specific OD evaluation.

PREDICTION MODEL / DECISION CRITERIA
OECD 431:
The cut-off value of 35% and classification method was validated in an international validation study of this kit (Fentem, 1998).
For 2 disks:
If both disks have mean viability of ≥ 35% = Non Corrosive
If both disks have mean viability of < 35% = Corrosive (at the corresponding incubation period)

For more than 2 disks:
If the mean value is ≥35% and the variability is less than 50% = Non Corrosive
If the mean value is <35% and the variability is less than 50% = Corrosive

Otherwise:
If the classification is not made with these criteria, retest with 2 more disks. Take the mean of the 4 disks to classify as above or below 35%. Outlier values may be excluded where there are scientific reasons, such as where application or rinsing is difficult and that the Study Director considers that a result is not representative.

OECD 439:
In the present study, the irritancy potential of test items is predicted by the mean tissue viability of tissues exposed to the test item. The test item is identified as requiring classification and labelling according to UN GHS (Category 2 or Category 1) if the mean relative viability of three individual tissues after 15 minutes exposure to the test item and 42 hours post incubation is less or equal (≤) to 50% of the mean viability of the negative controls. In case the test item is found to be non-corrosive, and shows tissue viability after exposure and post-treatment incubation is less than or equal (≤) to 50%, the test item is considered to be irritant to skin in accordance with UN GHS Category 2. The test item may be considered to be non-irritant to skin in accordance with UN GHS (No Category), if the mean relative viability of three individual tissues after 15 minutes exposure to the test item and 42 hours post incubation is more than (˃) to 50% of the mean viability of the negative controls.

Validity of the Test System:
General validity criteria:
- The mean OD value of the two or three negative control tissues should be ≥ 0.6 and ≤ 1.5
- Control OD values should be within the historical control range.
- The mean OD value of the blank samples (acidified isopropanol) should be < 0.1.
- Specific criteria for corrosivity testing:
The difference of viability between the two tissue replicates should not exceed 30%. The acceptable mean percentage viability range for positive controls is ≤ 20%.

Specific criteria for irritation testing:
Standard deviation value (SD) of the negative control tissues and the identically treated replicates % viability should be ≤ 18. The acceptable mean percentage viability range for positive controls is 0-40% and the standard deviation value (SD) of the % viability should be ≤ 18.

Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
Irritation test
- Amount(s) applied (volume or weight with unit): 20 µL
Corrosion test
- Amount(s) applied (volume or weight with unit): 50 µL

NEGATIVE CONTROL
Irritation test
- Amount(s) applied (volume or weight): 50 µL PBS
Corrosion test
- Amount(s) applied (volume or weight): 50 µL
- Concentration (if solution): 0.9% (w/v) NaCl solution))

POSITIVE CONTROL
Irritation test
- Amount(s) applied (volume or weight): 50 µL
- Concentration (if solution): 5% (w/v) SDS solution
Corrosion test
- Amount(s) applied (volume or weight): 50 µL glacial acetic acid
Duration of treatment / exposure:
Irritation test: 15 min
Corrosion test: 4 h
Duration of post-treatment incubation (if applicable):
42 h for irritation testing
Number of replicates:
- Two for corrosion test and two for irritation test
- Two negative controls and two positive controls were run in corrosivity testing and three negative controls and three positive controls were run in irritation testing . As the test item was coloured, two additional test item-treated living tissues were used for the non specific OD evaluation.

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
% tissue viability
Remarks:
OECD 431
Run / experiment:
mean of replicates
Value:
82.4
Vehicle controls validity:
not examined
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Irritation / corrosion parameter:
% tissue viability
Remarks:
OECD 439
Run / experiment:
mean of triplicates
Value:
97.2
Vehicle controls validity:
not examined
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
- OTHER EFFECTS:
- Visible damage on test system: None

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
In this study under the given conditions, the test item showed no irritant or corrosive effects. The relative mean tissue viability was > 50% for irritation and > 35 % for corrosivity testing. The test item is therefore classified as “non-irritant” and "non-corrosive" in accordance with UN GHS “No Category”.
Executive summary:

In a combined in vitro skin irritation/corrosion study conducted according to OECD guidelines 431 and 439, the skin corrosion and irritation potential of the test item, Ethoxylated phenol phosphate, was analysed by using the three-dimensional human skin model EpiSkin-SM (SkinEthic) comprising a reconstructed human epidermis with functional stratum corneum. The mean cell viability was above the threshold of 35% (82.4%) in the corrosivity test and above the threshold of 50% (97.2%) in the irritation test, as compared to the negative control. Based on these results, the test item is considered to be non - irritant and non – corrosive to the skin and no classification is warranted.