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)).

Endpoint summary

Description of key information

Based on the results of the available studies, the test material is not considered irritating to the eye or to the skin.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

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.

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

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

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).

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

other: 3 minutes

Score:

85.5

Reversibility:

other: no irritation effects seen

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

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

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.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experimental Start Date 06 Mar 2013 Experimental completion Date: 06 Mar 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.

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

other: not applicable as this is an in vitro study

Strain:

other: not applicable as this is an in vitro study

Details on test animals or tissues and environmental conditions:

Not applicable as this is an in vitro study. However the bovine eyes were received from JW. Treuth & Sons, Inc. on 06 Mar 2013 and transported overnight to MB Research in Hank's Balanced Salt Solution with Pennstrep in a refrigerated container.

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

Liquids were tested undiluted. Following the pretest observations, the MEM solution was removed from the anterior chamber. A volume of 0.75 ml of the ethanol, MEM or liquid test article was applied to the epithelium of each of the three (3) positive controls, three (3) negative controls, or three (3) test articletreated corneas in a manner, which ensured the entire cornea was covered.

Duration of treatment / exposure:

10 minutes

Observation period (in vivo):

2 hours

Number of animals or in vitro replicates:

Not applicable as an in vitro study was conducted

Details on study design:

All holders and corneas were placed in a horizontal position (anterior side up) in the 32°C (± 1°C) incubator. After 10.:!:. 1 minute, the test article, ethanol or MEM solution in the controls were removed from the epithelium of the cornea and anterior chamber of the holder by washing with MEM solution containing phenol red. The anterior and posterior chambers of the holders were refilled with fresh MEM solution. Opacity measurements were made following the 10-minute exposure and MEM solution refill.

All corneas were incubated at 32°C (± 1°C) for an additional two hours at which time the MEM solution in the anterior and posterior chambers was removed and the holders refilled with fresh MEM solution. A measurement of opacity was taken with each treated cornea compared to the blank supplied with the OPKIT. This is the reading that was used in the final in-vitro calculations.

Immediately following the 2-hour opacity measurement, the MEM solution was removed from the anterior chamber and replaced with 1.0 ml of 0.4% sodium fluorescein solution (in Dulbecco's Phosphate Buffered Saline) for liquid test articles and corresponding controls. Each holder was returned to the 32°C (± 1°C) incubator in a horizontal position (anterior side up) ensuring contact of the fluorescein with the cornea.

After 90 minutes, the fluid from the posterior chamber was removed and the amount of dye, which passed through the cornea, was measured as the optical density at 490 nm by a plate reader or spectrophotometer.

Irritation parameter:

other: calculated in vitro score

Basis:

mean

Time point:

other: 2 to 4 hours

Score:

1.43

Reversibility:

fully reversible

Remarks on result:

other: As the scpre is < 55.1 the test substance does not need to be classified

Irritant / corrosive response data:

Test material - In vitro Score: 1.43
Negative Control (MEM) - In vitro score: 0.71
Positive control (ethanol) - In vitro score: 43.27

Other effects:

The corrected mean opacity score was 1.34. The corrected mean optical density (permeability) score was 0.006.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Conclusion: The in vitro score was calculated as 1.43.

Executive summary:

Objective: To determine the potential for ocular irritation using an alternative to the Draize methodology. This protocol is based on the methodology described in the OECD Guideline for the Testing of Chemicals #437, adopted September 7,2009.

Method Synopsis: Three corneas were dosed with 0.75 ml of CAS# 1365345-64-7 (SE7B Batch 2137-0). Opacity measurements and sodium fluorescein permeability were determined.

Summary: The corrected mean opacity score was 1.34. Controls were within normal limits. The corrected mean optical density (permeability) score was 0.006.

Conclusion: The in vitro score was calculated as 1.43.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Justification for selection of skin irritation / corrosion endpoint:
Based on the results of this study, the test material is not corrosive to the skin.

Justification for selection of eye irritation endpoint:
Conclusion: The in vitro score was calculated as 1.43.

Justification for classification or non-classification

All available data indicates that the substance demonstrates no irritation/corrosion to the eyes or skin.