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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:
29 Aug 2017 - 31 Aug 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report date:
2018

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)
Deviations:
no
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
Tetrakis(2-ethylbutyl) orthosilicate
EC Number:
201-085-9
EC Name:
Tetrakis(2-ethylbutyl) orthosilicate
Cas Number:
78-13-7
Molecular formula:
C24H52O4Si
IUPAC Name:
tetrakis(2-ethylbutyl) orthosilicate
Test material form:
liquid
Details on test material:
Tetrakis(2-ethylbutyl) Orthosilicate
CAS 78-13-7
Expiration: May 2020
Clear colorless liquid
Specific details on test material used for the study:
Source: Gelest, Inc. (Morrisville, PA)
Lot #8E-33-134
Expiration date: May 2020

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: stored in dessicator; room temperature
- Solubility and stability of the test substance in the solvent/vehicle: not applicable

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: used as received

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Details on animal used as source of test system:
not applicable
Justification for test system used:
The EpiDerm™ Skin Model closely parallels human skin, and is recommended by test guidelines. Validation studies have reported that tests employing human skin models are able to reliably discriminate between known skin corrosives and non-corrosives.
Vehicle:
unchanged (no vehicle)
Details on test system:
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: MatTek Corporation (Ashland, MA) EpiDerm™ tissues
- Tissue batch number(s): Lot No. 26787 Kit I
- Shipping date: 28 August 2017
- Delivery date: 29 August 2017
- Date of initiation of testing: 30 August 2017

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature
- Temperature of post-treatment incubation (if applicable): not applicable

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: not specified
- Observable damage in the tissue due to washing: none reported
- Modifications to validated SOP: none

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL Dulbecco's Modified Eagle's Medium (DMEM)
- Incubation time: 3 hours
- Spectrophotometer: μQuant Plate Reader (Bio-Tek Instruments, Winooski, VT).
- Wavelength: 540 nM


FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: Tissue viability was assessed with an MTT QC assay. Application of tissue culture water to the tissues for four hours produced an MTT OD value of 1.7 for Lot No. 26787 Kit I. This value met the criteria of >1.0 but < 3.0 indicating adequate viability of tissues. The historical tissue viability values from the 1996 EpiDerm Database (N=184) for water are mean= 1.47; standard deviation = 0.13; Range ± 2 standar deviatons = 1.21 - 1.73.
- Barrier function: Tissues were exposed to 1% Triton X-100 for 4, 6, 8 and 10 hours. The time of exposure required to reduce the tissue viability (ET-50) using the MTT viability assay is determined (See MatTek EpiDerm MTT ET-50 Protocol) for each lot of tissue. ET-50’s must fall within the range of the 1996 EpiDerm database of 4.77 – 8.72 hours. The ET50 for Lot No. 26787 Kit I was 7.57 hours.
- Morphology: Tissue viability and barrier function tests are wtihin the acceptable ranges and indicate appropriate formation of the epidermal barrier, the presence of a function stratum corneum, a viable basal cell layer and intermediate spinous and granular layers.
- Contamination: The cells used to produced EpiDerm tissues are screened for potential biological contaminants by the manufacturer. Tests were performed by MatTek for each of the potential biological contaminants as follows: HIV (oligonucleotide-directed amplification), Hepatitis-B (oligonucleotide-directed amplification), Hepatitis-C (oligonucleotide-directed amplification), and Bacteria, yeast and other fungi (long-term antibiotic, antimycotic free culture)
- Reproducibility: Each lot of tissues was Quality Assured by MatTek according to specific QC standards including: histology, tissue viability (MTT mean optical density), reproducibility (SD) and tissue thickness. Tissues were used consistently on the same day, i.e. following overnight storage at 4°C (Wednesday morning) to maintain reproducibility.


NUMBER OF REPLICATE TISSUES: 2

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

DECISION CRITERIA
- 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 (volume or weight with unit): 50 microliters of test article was applied to the top of each tissue
- Concentration (if solution): not applicable

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 microliters of test article was applied to the top of each tissue
- Concentration (if solution): not applicable

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 50 microliters of test article was applied to the top of each tissue
- Concentration (if solution): 8N
Duration of treatment / exposure:
Each test article remained in contact with the EpiDerm™ tissue for 3 and 60 minutes.
Duration of post-treatment incubation (if applicable):
N/A
Number of replicates:
Duplicate EpiDerm™ tissues

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
60 minutes
Value:
104.7
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
3 minutes
Value:
91.7
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
Viability differences between the two identically treated tissues in all samples and controls at 3 minutes were 0.7% to 5.4%. Viability differences between the two identically treated tissues at 60 minutes were 0.8% to 8.9%. Inter-tissue viability differences at both time points met the acceptance criterion (<30%). Calibration of the uQuant microplate reader passed all acceptance criteria.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Skin corrosion is defined as the production of irreversible tissue damage in skin following the application of test material. The percent viability following a 3 minute and 60 minute exposure was used to determine corrosivity potential according to OECD TG 431. For mean viability ≥ 50% at 3 minutes and >15%, the test article is determined to be non-corrosive, and ≥ 50% at 3 minutes but <15% at 60 minutes, the test article is determined to be corrosive. In this case, the mean viability of Tetrakis(2-ethylbutyl) Orthosilicate was 91.7 % following a 3 minute exposure and 104.7% following a 60 minute exposure. This finding does not warrant classification of Tetrakis(2-ethyl butyl) Orthosilicate as a Category 1 skin corrosive substance under the Regulation (EC) 1272/2008 on classification, labeling, and packaging of substances and mixtures (CLP).
Executive summary:

The skin corrosive potential of Tetrakis (2 -ehtylbutyl) Orthosilicate was evaluated using the MatTek EpiDerm™ Skin Corrosivity Test (OECD TG 431). MatTek EpiDerm™ tissues were treated in duplicate with the test articles, negative control and positive control for 3 minutes and 60 minutes according. Following treatment, the viability of the tissues was determined using methyl thiazole tetrazolium (MTT) uptake and conversion, and the absorbance of each tissue was measured at 540 nm. The mean viability was then expressed as a percent of control values. The percent viability was used to determine corrosivity potential. The percent viability following a 3 minute and 60 minute exposure was used to determine corrosivity potential. For mean viability ≥ 50% at 3 minutes and >15%, the test article is determined to be non-corrosive, and ≥ 50% at 3 minutes but <15% at 60 minutes, the test article is determined to be corrosive. In this case, the mean viability of Tetrakis(2 -ethylbutyl) Orthosilicate was 91.7% following a 3 minute exposure and 104.7% following a 60 minute exposure. This finding does not warrant classification of Tetrakis(2 -ethylbutyl) Orthosilicate as a Category 1 skin corrosive substance under the  Regulation (EC) 1272/2008 on classification, labeling, and packaging of substances and mixtures (CLP).