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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin corrosion: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
August 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
4-(1-{3,5-Bis[(dimethylamino)methyl]-4-hydroxyphenyl}-1-methylethyl)-2,6-bis[(dimethylamino)methyl]phenol
Molecular formula:
C27H44N4O2
IUPAC Name:
4-(1-{3,5-Bis[(dimethylamino)methyl]-4-hydroxyphenyl}-1-methylethyl)-2,6-bis[(dimethylamino)methyl]phenol
Constituent 2
Chemical structure
Reference substance name:
2,6-Bis[(dimethylamino)methyl]-4-(1-{3-[(dimethylamino)methyl]-4-hydroxyphenyl}- 1-methylethyl)phenol
Molecular formula:
C24H37N3O2
IUPAC Name:
2,6-Bis[(dimethylamino)methyl]-4-(1-{3-[(dimethylamino)methyl]-4-hydroxyphenyl}- 1-methylethyl)phenol
Constituent 3
Chemical structure
Reference substance name:
2-[(Dimethylamino)methyl]-4-(1-{3-[(dimethylamino)methyl]-4-hydroxyphenyl}-1-methylethyl)phenol
Molecular formula:
C21H30N2O2
IUPAC Name:
2-[(Dimethylamino)methyl]-4-(1-{3-[(dimethylamino)methyl]-4-hydroxyphenyl}-1-methylethyl)phenol
impurity 1
Reference substance name:
Unknown impurities
IUPAC Name:
Unknown impurities
impurity 2
Chemical structure
Reference substance name:
Xylene
EC Number:
215-535-7
EC Name:
Xylene
Cas Number:
1330-20-7
Molecular formula:
C8H10
IUPAC Name:
Benzene, dimethyl-
impurity 3
Chemical structure
Reference substance name:
Water
EC Number:
231-791-2
EC Name:
Water
Cas Number:
7732-18-5
Molecular formula:
H2O
IUPAC Name:
water
impurity 4
Reference substance name:
Sulphated ash
IUPAC Name:
Sulphated ash
Test material form:
solid: bulk
Details on test material:
- Name of test material (as cited in study report): HPP 12879-1
Specific details on test material used for the study:
TREATMENT OF TEST MATERIAL PRIOR TO TESTING

- Treatment of test material prior to testing: test item was melted in a hot cabinet at 90 °C.

FORM AS APPLIED IN THE TEST (if different from that of starting material): liquid

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Justification for test system used:
commercially available test system
Vehicle:
unchanged (no vehicle)
Details on test system:
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: epiCS® (CellSystems, Troisdorf, Germany).
- Tissue batch number(s): Cat.-No.CS-1001

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature for 3 min- exposure and incubator (37 ± 2° C) for 60 min- exposure

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1mg/ml
- Incubation time: 3 hours
- Spectrophotometer: EL808, Bio-Tek
- Wavelength: 570 nm

NUMBER OF INDEPENDENT TESTING RUNS / EXPERIMENTS TO DERIVE FINAL PREDICTION:
the optical density of the isopropanol-extracts of 3 inserts was determined by duplicate per insert = 6 OD values.

PREDICTION MODEL / DECISION CRITERIA:
- The mean optical density (OD) value obtained with the test item was used to calculate the percentage of viability relative to the negative control, which is set at 100 %.
- classification according to UN GHS: if the mean percent tissue viability after 3 min-exposure is less than (<) 50 % the test item is classified as corrosive (Category 1A); if the mean percent tissue viability after 3-min-exposure is greater than or equal (≥) to 50 % AND < 15% after 60-min exposure the test item is classified as corrosive (Category 1B/1C); if the mean percent tissue viability after 60-min-exposure is ≥ 50 % after 3-min exposure AND ≥ 15% after 60-min exposure the test item is considered as non-corrosive

The corrosive potential of the test item is assessed by determination of its cytotoxic effect on an in vitro reconstructed human epidermis. The test principle is based on the MTT assay reflecting the cell viability after exposure to the topically applied test item.
All tests were performed in triplets for each time point. The test item was applied undiluted , i.e. 50 µl of the melted and cooled down test item per insert for 3 min. (room temperature) and 60 min. Cell viability was measured by the amount of MTT reduction (calculated on the basis of optical density of the negative control).
Control samples:
yes, concurrent negative control
Amount/concentration applied:
TEST MATERIAL
- Amount(s) applied: 50 μl

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 μl
- Concentration (if solution): 0.9% NaCl in water
Duration of treatment / exposure:
3 min and 60 min
Number of replicates:
3

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
exposure period: 3 min.
Value:
ca. 104.58
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
not specified
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
exposure period: 60 min.
Value:
ca. 71.42
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
not specified
Other effects / acceptance of results:
DEMONSTRATION OF TECHNICAL PROFICIENCY:
- Reliability of the test was previously confirmed by interlaboratory validation

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes
- Acceptance criteria met for positive control: yes, within regular intervals in the lab
- Acceptance criteria met for variability between replicate measurements: yes

Any other information on results incl. tables

Table 1: Tabular Summary of the results

 Sample No.  Test item  Expousre time [min]  OD mean*  StdDev  % Viability
 1 -3  negative control, 0.9% NaCl  60  2.34  0.38  100
 7 -9  HPP 12879 -1  60  1.67  0.22  71.42
 10 -12  negative control, 0.9% NaCl  3  2.21  0.39  100
 16 -18 HPP 12879 -1  3  2.31  0.20  104.58

*6 values

Applicant's summary and conclusion

Interpretation of results:
other: negative
Executive summary:

A study was performed for the assessment of the skin corrosion of the test item with reconstructed human epidermis (RhE). The experiment was carried out using the commercially available test method epiCS®. The study was conducted in accordance with OECD TG 431 and EU Test Method B.40 bis. The mean value of cell viability was recorded to be 105 % (3 min. exposure) and 71% (60 min. exposure). The test item was thus shown to be not corrosive to reconstructed human skin in vitro.

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