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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:
key study
Study period:
24 Oct 2016 - 28 Oct 2016
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 guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)
Version / remarks:
29 July 2016
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: EU method B.40 BIS (In Vitro Skin corrosion: Human Skin Model Test), adopted 31 May 2008
Version / remarks:
31 May 2008
Deviations:
no
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
Tributyl-2-thiourea
EC Number:
219-350-2
EC Name:
Tributyl-2-thiourea
Cas Number:
2422-88-0
Molecular formula:
C13H28N2S
IUPAC Name:
1,1,3-tributylthiourea
Test material form:
liquid
Details on test material:
- Name of test material (as cited in study report): Tri butyl thiourea
- Appearance: Clear pale to medium brown liquid
- Storage conditions: At room temperature

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Cell source:
skin obtained from plastic surgery from multiple donors
Details on animal used as source of test system:
EpiDerm™ Reconstructed Human Epidermis
- All cells used to produce Eipderm™ are purchased or derived from tissue obtained by MatTek Corporation from acredited institutions.
- Cells are screened for potential biological contaminants (HIV-1, Hepatitis B, Hepatitis C, bacteria, yeast and fungi)
Justification for test system used:
Recommended test system in international guidelines (OECD and EC).
Details on test system:
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm Skin Model (EPI-200)
- Tissue lot number: 24900 Kit K and J

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: All incubations, with the exception of the test item incubation of 3 minutes at room temperature, were carried out in a controlled environment at 37.0 ± 1.0°C.

REMOVAL OF TEST MATERIAL AND CONTROLS
- After the exposure period, the tissues were washed with phosphate buffered saline to remove residual test item.

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 5 mg/mL
- Incubation time: 3 h
- Spectrophotometer: TECAN Infinite® M200 Pro Plate Reader
- Wavelength: 570 nm

NUMBER OF REPLICATE TISSUES: 2 replicates per exposure duration, one negative control, one positive contol.

ACCEPTANCE OF RESULTS:
The in vitro skin corrosion test is considered acceptable if it meets the following criteria:
a) The absolute mean OD570 of the two tissues of the negative control should reasonably be within the laboratory historical control data range.
b) The mean relative tissue viability following 3-minute exposure to the positive control should be ≤ 30%.
c) In the range of 20 – 100% viability, the maximum inter-tissue variability (in viability) is ≤ 30% between two tissues treated identically.
d) In the range of 20 – 100% viability, the maximum difference in percentage between the mean viability of two tissues and one of the two tissues is ≤ 15%.

DECISION CRITERIA
A test item is considered corrosive in the skin corrosion test if:
a) The relative mean tissue viability obtained after 3-minute treatment compared to the negative control tissues is decreased below 50%.
b) In addition, a test item considered non-corrosive (viability ≥ 50%) after the 3-minute treatment is considered corrosive if the relative tissue viability after 1-hour treatment with the test item is decreased below 15%.

A test item is considered non corrosive in the in vitro skin corrosion test if:
a) The relative mean tissue viability obtained after the 3-minute treatment compared to the negative control tissues is not decreased below 50%.
b) In addition, the relative tissue viability after the 1-hour treatment is not decreased below 15%.
Control samples:
yes, concurrent negative control
yes, concurrent positive control
yes, concurrent MTT non-specific colour control
other: concurrent control for MTT reduction by test item
Amount/concentration applied:
50 μL
Duration of treatment / exposure:
3-minute and 1-hour
Duration of post-treatment incubation (if applicable):
none

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
3-minute exposure
Value:
107
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Remarks:
(100%)
Positive controls validity:
valid
Remarks:
(7%)
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
1-hour exposure
Value:
107
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Remarks:
(100%)
Positive controls validity:
valid
Remarks:
(7%)
Other effects / acceptance of results:
- OTHER EFFECTS:
- Direct-MTT reduction: No
- Colour interference with MTT: No

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes, mean relative tissue viability following 3-minute exposure was 7%.
- Acceptance criteria met for variability between replicate measurements: Yes

Table 1 shows the results of individual OD measurements at 570 nm.

Any other information on results incl. tables

Table 1 Individual OD measurements at 570 nm.

 

3-minute application (OD570)

1-hour application (OD570)

 

A

B

A

B

Negative control

Measurement 1

Measurement 2

Measurement 3

 

1.4673

1.4841

1.5029

 

1.4019

1.4786

1.4655

 

1.6350

1.5955

1.6174

 

1.5940

1.5358

1.5578

Tri butyl thiourea

Measurement 1

Measurement 2

Measurement 3

 

1.6784

1.6460

1.7092

 

1.4655

1.4503

1.4801

 

1.8208

1.7385

1.7546

 

1.6272

1.5979

1.6275

Positive control

Measurement 1

Measurement 2

Measurement 3

 

0.1538

0.1517

0.1549

 

0.1348

0.1357

0.1354

 

0.1320

0.1297

0.1318

 

0.1579

0.1651

0.1579

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
An in vitro skin corrosion test was conducted with Tri butyl thiourea according to OECD 431 guideline and GLP principles.
It is concluded that this test is valid and that the test substance is not corrosive in the in vitro skin corrosion test.
Executive summary:

In an in vitro skin corrosion test using a human skin model ( EpiDerm Skin Model, Lot no.: 24900 Kit K and J), the influence of the test substance on the viability of human skin was tested. Fifty μL of Tri butyl thiourea was applied directly to 0.6 cm2 cultured skin. After 3-minute and 1-hour treatments the substance was removed and the viability of the cells was tested by reduction of MTT. Viability of unexposed skin was set at 100%. The relative mean tissue viability obtained after 3-minute and 1-hour treatments with test substance compared to the negative control tissues was 107% for both exposure times. The positive control had a mean cell viability of 7% after 3 minutes exposure.

Since the mean relative tissue viability was not below 50% after the 3-minute treatment and not below 15% after the 1-hour treatment, it can be concluded that the test substance is not corrosive in the in vitro skin corrosion test.