2-Propenoic acid, 2-methyl-, tetracosyl ester, branched

Endpoint summary

Description of key information

The in vitro skin irritation of 2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) was determined in accordance with the OECD Guideline for Testing of Chemicals 439 during a GLP-compliant study. This in vitro risk assessment assay predicts the Skin irritation potential of a chemical by measurement of its cytotoxic effect on the EpiDerm™ tissue model.

Prior to testing, the test substance was checked for interference with water and/or MTT. No interference was identified.

Skin irritation of the test substance and controls was evaluated in triplicate. After 60±1 minutes exposure on the surface of the EpiDerm™ reconstructed human epidermis and a 42±4h post-exposure incubation time, viability of the tissues was assessed and compared to the negative control.

The percentage of viability obtained with 2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) was 128.113%, therefore it was considered as non-irritant to the skin according to the criteria laid down in the OECD Guideline for Testing of Chemicals 439.

The substance was not expected to be corrosive to the skin, therefore an in vitro skin irritation study was conducted in a first instance. The in vitro skin irritation study was sufficient to conclude on the classification of the substance for this endpoint, therefore, it was not required to conduct an in vitro skin corrosion study, in accordance with Annex VII, Column 2, of REACH.

An in vitro eye irritation study was performed in line with OECD Testing Guideline 437 (Bovine Corneal Opacity and Permeability Test Method). The study was GLP-compliant.

The undiluted test material was applied to three cattle corneas obtained from an abattoir at a volume of 0.75 mL, after which each cornea was incubated at 32±1°C for 10 minutes. Opacity and permeability of corneas were measured following exposition.

Negative and positive controls were performed and considered as valid.

An In Vitro Irritation Score (IVIS) of 0.8 was calculated for 2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) from corneal opacity and permeability measurements. The substance is not irritant to the eye according to the guideline.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 15 November 2017 to 23 January 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm (EPI-200) Reconstituted Human Epidermis
- Tissue batch number(s): 25864

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37°C

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: Optical Density (O.D.) values: 1.601 ± 0.007 (mean and SD of MTT value of 3 tissues exposed to H2O). Acceptance range: [1.0 - 3.0]- PASS
- Barrier function: ET 50% of tissues exposed to 100 µL Triton X-100 1% (n=3): 5.84 h. Acceptance range: [4.77h- 8.72h] - PASS
- Morphology: Tissue thickness: 128 µm. Acceptance range: >70 µm and <130 µm - PASS
- Contamination: No contamination reported - PASS

NUMBER OF REPLICATE TISSUES:
Triplicate

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 30 μL neat substance
NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 μL
POSITIVE CONTROL
- Amount(s) applied (volume or weight): 30 μL

Duration of treatment / exposure:

60 ± 1 minute (25 minutes at room temperature and 35 minutes at 37°C, 5% CO2, ≥95% RH)

Duration of post-treatment incubation (if applicable):

42 ± 4h post-treatment incubation (at 37°C, 5% CO2, ≥ 95% RH)

Number of replicates:

Triplicate

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Mean value

Value:

128.113

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: None reported
- Direct-MTT reduction: No direct reduction
- Colour interference with MTT: No colour interference

Interpretation of results:

GHS criteria not met

Conclusions:

2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) did not reduce the tissue viability. The test substance does not meet the criteria for classification as irritant to the skin according to Regulation (EC) No.1272/2008.

Executive summary:

The in vitro skin irritation of 2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) was determined in accordance with the OECD Guideline for Testing of Chemicals 439 during a GLP-compliant study. This in vitro risk assessment assay predicts the Skin irritation potential of a chemical by measurement of its cytotoxic effect on the EpiDerm™ tissue model.

Prior to testing, the test substance was checked for interference with water and/or MTT. No interference was identified.

Skin irritation of the test substance and controls was evaluated in triplicate. After 60±1 minutes exposure on the surface of the EpiDerm™ reconstructed human epidermis and a 42±4h post-exposure incubation time, viability of the tissues was assessed and compared to the negative control.

The percentage of viability obtained with 2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) was 128.113%, therefore it was considered as non-irritant to the skin according to the criteria laid down in the OECD Guideline for Testing of Chemicals 439.

The test item did not meet the criteria for classification as irritant to the skin according to Regulation (EC) No.1272/2008 on the Classification, Labelling and Packaging of Substances and Mixtures.

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:

From 22 February 2018 to 26 March 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU method B.47 (Bovine corneal opacity and permeability test method for identifying ocular corrosives and severe irritants)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

Eyes from adult cattle (typically 12 to 60 months old) were obtained from a local abattoir as a by-product from freshly slaughtered animals. The eyes were excised by an abattoir employee after slaughter, and were placed in Hanks’ Balanced Salt Solution (HBSS) supplemented with antibiotics (penicillin at 100 IU/mL and streptomycin at 100 μg/mL). They were transported to the test facility over ice packs on the same day of slaughter. The corneas were prepared immediately on arrival.

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.75 mL neat substance

Duration of treatment / exposure:

10 minutes

Duration of post- treatment incubation (in vitro):

120 minutes

Number of animals or in vitro replicates:

Three corneas

Details on study design:

SELECTION AND PREPARATION OF CORNEAS
The medium from both chambers of each holder was replaced with fresh complete EMEM. A pre-treatment opacity reading was taken for each cornea using a calibrated opacitometer. The average opacity for all corneas was calculated. Three corneas were randomly allocated to the negative control. Three corneas were also allocated to the test item and three corneas to the positive control item.

NUMBER OF REPLICATES
Triplicate

NEGATIVE CONTROL USED
Sodium chloride 0.9% w/v

POSITIVE CONTROL USED
Ethanol

APPLICATION DOSE AND EXPOSURE TIME
A volume of 750 μL of the test article was applied to each of three corneas followed by a 10 minute incubation at 32 °C.

TREATMENT METHOD: [closed chamber / open chamber]

POST-INCUBATION PERIOD: yes

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: 3
- POST-EXPOSURE INCUBATION: 120 minutes

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: Labtech LT-4500 microplate reader
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of microtiter plate reader (OD492)

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

Irritation parameter:

in vitro irritation score

Value:

0.8

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: The corneas treated with the test item were clear post treatment and post incubation.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The negative control gave opacity of ≤3.0 and permeability ≤0.077. The negative control acceptance criteria were therefore satisfied.
- Acceptance criteria met for positive control: The positive control In Vitro Irritancy Score was within the range of 31.6 to 58.7. The positive control acceptance criterion was therefore satisfied.

Interpretation of results:

GHS criteria not met

Conclusions:

2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) produced an IVIS of 0.8 during an in vitro eye irritation study. The test substance does not meet the criteria for classification as irritant to the eye according to Regulation (EC) No.1272/2008.

Executive summary:

An in vitro Bovine Corneal Opacity and Permeability (BCOP) assay was performed in line with OECD

Testing Guideline 437 (Bovine Corneal Opacity and Permeability Test Method). The study was GLP-compliant.

The undiluted test material was applied to three cattle corneas obtained from an abattoir at a volume of 0.75 mL, after which each cornea was incubated at 32±1°C for 10 minutes. Opacity and permeability of corneas were measured following exposition.

Negative and positive controls were performed and considered as valid.

An In Vitro Irritation Score (IVIS) of 0.8 was calculated for 2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) from corneal opacity and permeability measurements. The substance is not irritant to the eye according to the guideline.

It can be concluded that 2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) does not meet the criteria for classification as irritant to the eye in accordance with Regulation (EC) No.1272/2008.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Justification for classification or non-classification

Skin irritation and eye irritation were investigated during GLP-compliant studies performed according to relevant OECD Guidelines.

2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) did not reduce the tissue viability during an in vitro skin irritation study. The test substance does not meet the criteria for classification as irritant to the skin according to Regulation (EC) No.1272/2008.

2-Propenoic acid, 2-methyl-, tetracosyl ester(branched) produced an IVIS of 0.8 during an in vitro eye irritation study. The test substance does not meet the criteria for classification as irritant to the eye according to Regulation (EC) No.1272/2008.