Registration Dossier

Administrative data

Description of key information

Key value for chemical safety assessment

Skin irritation / corrosion

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:
29th of May 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: guideline study
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test material information:
Composition 1
Details on test animals or tissues and environmental conditions:
Eyes from adult cattle were obtained from a local abattoir as a by-product from freshly slaughtered animals. The eyes were excised by an abattoir employee and placed in Hanks' Balances Salt Solution (HBSS), supplemented with Penicillin/Streptomycin, and transported to the laboratory on ice packs. The corneas were prepared immediately on arrival.
Vehicle:
unchanged (no vehicle)
Controls:
other: not applicable (control corneas were treated with the negative control)
Amount / concentration applied:
Each three corneae were exposed to 0.75 mL of the test item, the negative control (0.9% w/v sodium chloride solution), or the positive control (ethanol) for 10 minutes.
Duration of treatment / exposure:
Each holder was incubated, anterior chamber uppermost, at 32 ± 1 °C for 10 minutes.
Observation period (in vivo):
Not applicable
Number of animals or in vitro replicates:
Not applicable (three corneas were used for each treated series)
Details on study design:
After the application of the test item and control items the holders were gently titled back and forth to ensure a uniform application of the test item over the entire cornea. Each holder was incubated, anterior chamber uppermost, at 32 ± 1 °C for 10 minutes.

At the end of the exposure period the test item and control items were removed from the anterior chamber and the cornea was rinsed three times with fresh complete minimum essential medium (MEM) containing phenol red before a final rinse with complete MEM. The anterior chamber was refilled with fresh complete MEM. A post-treatment opacity reading was taken and each cornea was visually observed.

The holders were incubated, anterior chamber facing forward, at 32 ± 1 °C for 120 minutes.

After incubation the holders were removed from the incubator and a final opacity reading was taken. Each cornea was visually observed.

Following the opacity measurement the permeability of the corneas to soidum fluorescein was evaluated. The medium from the anterior chamber was removed and replaced with 1 mL of sodium fluorescein solution (4 mg/mL). The dosing holes were plugged and the holders incubated, anterior chamber uppermost, at 32 ± 1 °C for 90 minutes.

After the incubation in the posterior chamber each holder was decanted and retained.

360 µL of medium reoresenting each cornea was applied to a designated well on a 96 well plate and the optical density at 492 nm was measured.

If values greater than 1.500 OD492 were obtained a 1 in 5 dilution of the medium in complete MEM was performed and the measurement repeated. The modified value was multiplied by 5 to reflect the 1 in 5 dilution.
Irritation parameter:
other: In Vitro Irritancy Score Test Item
Basis:
mean
Score:
14.8
Reversibility:
other: not applicable
Irritation parameter:
other: In Vitro Negative Control
Basis:
mean
Score:
8.9
Reversibility:
other: not applicable
Irritation parameter:
other: In Vitro Irritancy Score Positive Control
Basis:
mean
Score:
31.8
Reversibility:
other: not applicable

Individual and mean corneal opacity measurements and individual and mean corneal permeability measurements are given in table 1 attached.

The condition of each cornea post treatment and at the final opacity measurement is given in table 2 attached.

The corneas treated with the test item were clear post treatment and cloudy post incubation. The corneas treated with the test negative control item were clear post treatment and post incubation. The corneas treated with the positive control item were cloudy post treatment and post incubation.

Interpretation of results:
other: The test item was considered not to be an ocular corrosive or sever irritant
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
The test item was considered not to be an ocular corrosive or severe irritant.
Executive summary:

A study was performed to assess the ocular irritancy potential of the test item to the isolated bovine cornea.

The undiluted test item was applied for 10 minutes followed for an incubation period of 120 mintes. Negative and positive controls ites were tested concurrently. The two endpoints, decreased light transmission through the cornea (opacity) and increased passage of sodium fluorescein dye through the cornea (permeability), were combined in an empirically derived formula to generate an In Vitro Irritancy Score (IVIS).

Interpretation:

A test item that induces an In Vitro Irritancy Score ≥ 55.1is defined as an ocular corrosive or sever irritant.

Results. The In Vitro irritancy scores are summarized as follows:

 Treatment In Vitro Irritancy Score 
 Test Item 14.8 
 Negative control 8.9 
 Positive Control 31.8 

The test item was considered not to be an ocular corroseve or severe irritant.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Additional information

The dermal irritant/ corrosive potential of the substance was assessed using:

- An in vitro skin corrosion test performed in the EPISKIN Reconstructed Human Epidermins Model according to OECD guideline 431 and Good Laboratory Practices (report number 41300981; endpoint name SK01 - In Vitro Skin Corr. EpiSkin (OECD 431, GLP))

- An in vitro Skin Irritation Potential Using the EPISKIN™ Resconstructed Human Epidermis Model according to OECD guideline 439 and EU method B.46. the study was performed in compliance with the Good Laboratory Practices (report number 41300980; SK02 - In Vitro Skin Irrit. EpiSkin)

The ocular irritant potential of the substance was assessed using:

-An in vitro Determination of Eye Irritation Potential using the Bovine Corneal Opacity and Permeability (BCOP) Assay performed according to OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants).

- An experimental study in a structurally similar substance (2,6 -Dimethyl-4 -heptanol CAS:108 -82 -7) in 5 rabbits based on the method published in "Draize, J.H., Woodard, G. AND Calvery, H.O.¨Methods for the study of irritation and toxicity of substances applied topically to the skin and mucous membranes. J. Pharmacol. & Exper. Therap., 82: 377, 1944." showing slight irritation potential.

- An experimental study in a structurally similar substance (2,6 -Dimethyl-4 -heptanol CAS:108 -82 -7) in 1 rabbits without guideline specified that showing moderate irritation but reversible effects in 72 hours.


Effects on eye irritation: moderately irritating

Justification for classification or non-classification

Skin irritation/corrosion:

Based on the results of the Guest (1987,1991) studies and according to the criteria laid down inEU directive 67/548/EEC andCLP (Reg. n° 1272/2008/EC), the substance is not classified on skin irritation.

Eye irritation:

Based on the result of the Gerbeix study (2010b) and according to the criteria laid down in EC regulation 1272/2008/EC and EU directive 67/548/EEC, the substance is classified as eye irritant category 2, H319: Causes serious eye irritation and R36 Irritating to eyes.