hydrogenation products of (esterification products of 2-ethylhexan-1-ol with (Estolide formation products of oleic acid and Fatty acids, C8-18 and C18-unsatd. (branched or linear)).

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation / corrosion

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Study conducted between 21st February 2013 and 28th March 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of relevant results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

hydrogenation products of (esterification products of 2-ethylhexan-1-ol with (Estolide formation products of oleic acid and Fatty acids, C8-18 and C18-unsatd. (branched or linear)).

EC Number:

696-271-3

Molecular formula:

n/a as UVCB

IUPAC Name:

hydrogenation products of (esterification products of 2-ethylhexan-1-ol with (Estolide formation products of oleic acid and Fatty acids, C8-18 and C18-unsatd. (branched or linear)).

Test animals

Species:

other: in vitro study therefore not applicable

Strain:

other: in vitro study therefore not applicable

Details on test animals or test system and environmental conditions:

in vitro study therefore not applicable

Test system

Type of coverage:

other: not applicable as in vitro study

Preparation of test site:

other: not applicable as in vitro study was conducted

Vehicle:

unchanged (no vehicle)

Controls:

other: A Negative Control (TCH20) and a Positive Control (KOH) were tested at 3 and 60 minutes, with a nylon mesh. Each treatment with test article or control was conducted in duplicate.

Amount / concentration applied:

50 µl of the test article were applied to the top of each EpiDerm tissue. The nylon mesh was then placed on top to facilitate even distribution of the test material. The test article remained in contact with the EpiDerm tissue for 3 and 60 minutes.

Duration of treatment / exposure:

The test article remained in contact with the EpiDerm tissue for 3 and 60 minutes.

Observation period:

At the end of the exposure period, each EpiDerm™ tissue was rinsed with phosphate buffered saline (PBS) and transferred to a 24-well plate containing 300 µI of MIT solution (1 mg/ml MIT in OMEM). The tissues were then returned to the incubator for a three-hour MTT incubation period. Following the MIT incubation period, each EpiDerm tissue was rinsed with PBS and then treated overnight with 2.0 ml of extractant solution (isopropanol) per well. The absorbency of an aliquot of the extracted MIT formazan was measured in triplicate at 540 nm using a microplate reader (µQuant Plate Reader, BioTek Instruments, Winooski, VT).

Number of animals:

Not applicable as an in vitro study was conducted

Details on study design:

EpiDerm™ Tissue Samples:
EpiDerm tissues, Lot 18161 Kit C, were received from MatTek Corporation (Ashland, MA) on 05 Mar 2013 and refrigerated at approximately 4°C. Before use, tissues were incubated (37°C ± 1°C, 5% ± 1% CO2) with assay medium (MatTek) for a one-hour equilibration. Equilibration medium was replaced with fresh medium before dosing.

Test Article Reduction of MTT:
100 µI of the test article were mixed with 1 ml of MIT solution (1 mg/ml Methyl thiazole tetrazolium diluted in Dulbecco's Modified Eagle's Medium (DMEM)). A Negative Control, 100 µI of tissue culture water, was tested concurrently. The solutions were incubated at room temperature in the dark for 60 minutes. After incubation, the solutions were visually inspected for purple coloration, which indicates that the test article reduced MIT. Since tissue viability is based on MIT reduction, direct reduction by a test article can exaggerate viability, making a test article seem less irritating than it really is. The test article did not reduce MIT and the assay continued as per the protocol.

Mesh Compatability:
A pre-cut nylon mesh supplied with the tissues was placed on a slide and 30 µI of the test article or tissue culture water was applied. After 60 minutes exposure, the mesh was checked microscopically. No interaction between the test article or tissue culture water and the mesh was observed so the test article was dosed using the mesh as a spreading aid.

Dosing:
50 µl of the test article were applied to the top of each EpiDerm tissue. The nylon mesh was then placed on top to facilitate even distribution of the test material. The test article remained in contact with the EpiDerm tissue for 3 and 60 minutes. A Negative Control (TCH20) and a Positive Control (KOH) were tested at 3 and 60 minutes, with a nylon mesh. Each treatment with test article or control was conducted in duplicate.

Tissue Viability (MTT Reduction):
At the end of the exposure period, each EpiDerm™ tissue was rinsed with phosphate buffered saline (PBS) and transferred to a 24-well plate containing 300 µI of MIT solution (1 mg/ml MIT in OMEM). The tissues were then returned to the incubator for a three-hour MTT incubation period. Following the MIT incubation period, each EpiDerm tissue was rinsed with PBS and then treated overnight with 2.0 ml of extractant solution (isopropanol) per well. The absorbency of an aliquot of the extracted MIT formazan was measured in triplicate at 540 nm using a microplate reader (µQuant Plate Reader, BioTek Instruments, Winooski, VT).

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

other: 3 minutes

Score:

85.5

Reversibility:

other: no irritation effects seen

Results 2

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

other: 60

Score:

98.8

Reversibility:

other: no irritation effects seen

Irritant / corrosive response data:

Quality Controls:
The mean OD of the Negative Control tissues was 2.212 at 3 minutes and 2.065 at 60 minutes, which met the acceptance criterion.

The mean relative tissue viability of the 60-minute Positive Control was 3.8%, which met the acceptance criterion.

Viability differences between the two identically treated tissues in all samples and controls at 3 minutes were 0.5% to 4.5%. Viability differences between the two identically treated tissues at 60 minutes were 0.3% to 9.3%. Inter-tissue viability differences at both time points met the acceptance criterion.

Other effects:

No other effects were noted in the study report

Any other information on results incl. tables

Test Article Identity	viability (3 min)	viability (60 min)	Predicted corrosivity
CAS# 1365345-64-7 (SE7B Batch 2137-0)	85.5 %	98.8 %	Non - corrosive
Tissue culture water (Negative Control)	100.0 %	100.0%	Non - corrosive
Potassium Hydroxide, 8.0 N (Positive Control)	22.2 %	3.8%	Corrosive

Applicant's summary and conclusion

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Based on the results of this study, the test material is not corrosive to the skin.

Executive summary:

OBJECTIVE: To predict and classify the skin corrosivity potential of a chemical by using a three dimensional human epidermis model. This protocol follows the procedures outlined in OECD Test Guideline 431, effective April 2004. METHOD SYNOPSIS: MatTek EpiDerm™ tissue samples were treated in duplicate with the test article, Negative Control and Positive Control for 3 minutes and 60 minutes. Following treatment, the viability of the tissues was determined using Methyl thiazole tetrazolium (MTT) uptake and conversion, and the absorbance of each sample was measured at 540 nm. The viability was then expressed as a percent of control values. The percent viability was used to determine corrosivity potential.

SUMMARY/CONCLUSION:

The results of the study can be seen below:

Test Article Identity	viability (3 min)	viability (60 min)	Predicted corrosivity
CAS# 1365345-64-7 (SE7B Batch 2137-0)	85.5 %	98.8 %	Non - corrosive
Tissue culture water (Negative Control)	100.0 %	100.0%	Non - corrosive
Potassium Hydroxide, 8.0 N (Positive Control)	22.2 %	3.8%	Corrosive

Based on the results of the study the test material is not corrosive. The test material will not be classified as corrosive, according to the Dangerous Substances Directive or to CLP.