Reaction mass of 1,2,3,4,4a,5,6,7-octahydro-2,5,5-trimethyl-2-naphthol and 4-(2,6,6-trimethyl-2-cyclohexen-1-yl)butan-2-one

Endpoint summary

Description of key information

Skin irritation (OECD 404): irritating

Eye irritation (OECD 438): non-corrosive

Eye irritation (OECD 492): irritating potential

Overall conclusion: irritating

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

28 May - 18 June 1991

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

No data on analytical purity of test substance given; extended to investigate 5 different concentrations of the test substance and a vehicle control on each animal

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

1981

Deviations:

yes

Remarks:

No data on analytical purity of test substance given; extended to investigate 5 different concentrations of the test substance and a vehicle control on each animal

GLP compliance:

yes (incl. QA statement)

Remarks:

The Danish National Testing Board, Denmark

Species:

rabbit

Strain:

other: Mol:Russian

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight at study initiation: 1.7 - 2.2 kg
- Housing: individually in sigle polypropylene cages (45x 55 cm) with perforated floor
- Diet: pelleted Altromin 2123 diet, ad libitum
- Water: ad libitum
- Acclimation period: one week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 3
- Humidity (%): 55 ± 15
- Air changes (per hr): 6
- Photoperiod (hrs dark / hrs light): 12 / 12

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

other: for test substance concentrations of 1, 10, 25 and 50% ethanol/diethylphthalat (1:1) was used as vehicle

Controls:

yes, concurrent vehicle

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.5 mL
- Concentrations: 1, 5, 10, 25% (w/w) and 100%

VEHICLE
- Amount applied: 0.5 mL

Duration of treatment / exposure:

4 h

Observation period:

21 days

Number of animals:

4

Details on study design:

TEST SITE
- Area of exposure: 10 x 10 cm on the dorsal area of the trunk divided into 6 test sites with a 1 cm wide adhesive tape: two anterior located test sites, two centrally located test sites and two posterior located test sites
- Type of wrap if used: The test substance or vehicle was applied to 6 gauze patches (2.5 x 2.5 cm) each and the patches were placed on the appropriate test sites on the back of each rabbit. The gauze patches were secured with a cross of 1 cm wide adhesive tape and fixed with Scanpor tape, 7.5 cm width, loosely wound rund the trunk.

REMOVAL OF TEST SUBSTANCE
- Washing: Treated skin was cleaned with soap and lukewarm water.
- Time after start of exposure: 4 h

OBSERVATION TIME POINTS
4.5, 24, 48 and 72 h after exposure and on Day 7, 14 and 21

SCORING SYSTEM:
- Method of calculation: Draize scoring system

Irritation parameter:

erythema score

Basis:

animal: #1, #2 and #4

Remarks:

100% test substance

Time point:

24/48/72 h

Score:

2.67

Max. score:

4

Reversibility:

not fully reversible within: 14 days

Remarks:

A dense layer of white scales was observed in all animals on Day 14; animal #1 and #4: scattered white scales with intact skin underneath on Day 21, indicating reversibility of effects; animal #2: no skin reaction on Day 21

Irritation parameter:

erythema score

Basis:

animal #3

Remarks:

100% test substance

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

not fully reversible within: 14 days

Remarks:

A dense layer of white scales was observed in all animals on Day 14; scattered white scales with intact skin underneath on Day 21, indicating reversibility of effects

Irritation parameter:

edema score

Basis:

animal #1

Remarks:

100% test substance

Time point:

24/48/72 h

Score:

2.67

Max. score:

4

Reversibility:

not specified

Irritation parameter:

edema score

Basis:

animal: #2 and #3

Remarks:

100% test substance

Time point:

24/48/72 h

Score:

1.67

Max. score:

4

Reversibility:

not specified

Irritation parameter:

edema score

Basis:

animal #4

Remarks:

100% test substance

Time point:

24/48/72 h

Score:

2.33

Max. score:

4

Reversibility:

not specified

Irritant / corrosive response data:

On day 7: A layer of scales was observed at the test sites in two rabbits treated with a 100% concentration (#1 and #2) and crusts were observed at the test sites in two rabbits treated with a 100% concentration (#3 and #4). A layer of scales was observed in all the rabbits at the test sites treated with a 25% concentration.
On day 14: A dense layer of white scales was observed at the test sites treated with a 100% concentration in all the rabbits. In rabbits #3 and #4 scattered white scales were observed at the test sites treated with a 25% concentration.
On day 21: Scattered white scales with intact skin underneath were observed at the test sites treated with a 100% concentration in three rabbits (#1, #3 and #4), in rabbit #2 no reaction on the skin was observed.

Table 1. Results of skin irritation study

Rabbit No	Location of test side	Test substance concentrations	Erythema-Eschar observed at hours					Edema observed at hours
			4.5	24	48	72	Mean 24/48/72 h	4.5	24	48	72	Mean 24/48/72 h
#1	A.L.	100%	0	2	3	3	2.67	1	2	3	3	2.67
	A.R.	25%	0	2	2	2	2.00	1	2	2	2	2.00
	M.L.	10%	0	0	0	0	0.00	0	0	0	0	0.00
	M.R.	5%	0	0	0	0	0.00	0	0	0	0	0.00
	P.L.	1%	0	0	0	0	0.00	0	0	0	0	0.00
	P.R.	V	0	0	0	0	0.00	0	0	0	0	0.00
#2	A.L.	V	0	0	0	0	0.00	0	0	0	0	0.00
	A.R.	5%	0	0	0	0	0.00	0	0	0	0	0.00
	M.L.	25%	0	2	2	2	2.00	0	1	1	1	1.00
	M.R.	1%	0	0	0	0	0.00	0	0	0	0	0.00
	P.L.	10%	0	0	0	0	0.00	0	0	0	0	0.00
	P.R.	100%	0	2	3	3	2.67	0	1	2	2	1.67
#3	A.L.	V	0	0	0	0	0.00	0	0	0	0	0.00
	A.R.	10%	0	1	1	1	1.00	0	1	1	0	0.67
	M.L.	100%	1	3	3	3	3.00	1	1	2	2	1.67
	M.R.	25%	1	2	2	2	2.00	1	2	2	2	2.00
	P.L.	5%	0	0	0	1	0.33	0	0	0	0	0.00
	P.R.	1%	0	1	0	0	0.33	0	0	0	0	0.00
#4	A.L.	100%	0	2	3	3	2.67	0	1	3	3	2.33
	A.R.	5%	0	0	0	0	0.00	0	0	0	0	0.00
	M.L.	1%	0	0	0	0	0.00	0	0	0	0	0.00
	M.R.	25%	0	2	2	2	2.00	0	1	1	1	1.00
	P.L.	10%	0	0	0	0	0.00	0	0	0	0	0.00
	P.R.	V	0	0	1	1	0.67	0	0	0	0	0.00

A.L.: anterior left treatment site

P.L.: posterior left treatment site

A.R.: anterior right treatment site

P.R.: posterior right treatment site

M.L.: mid-left treatment site

M.R.: mid-right treatment site

V: vehicle (ethanol/diethylphthalat (1:1))

Interpretation of results:

other: Skin Irrit. 2 according to Regulation (EC) No 1272/2008

Conclusions:

Under the conditions of this skin irritation study in rabbits the undiluted test substance (100%) and a concentration of 25% in ethanol/diethylphthalat (1:1) was irritating to the skin.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 Apr 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 438 (Isolated Chicken Eye Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)

Version / remarks:

adopted 26 Jul 2013

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Medicines & Healthcare products Regulatory Agency, UK

Species:

chicken

Strain:

not specified

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Source: Baileys Turkeys Ltd., Cheshire, UK
- Characteristics of donor animals: Chickens were approx. 56 days old and weighed 3 kg prior to slaughter.
- Storage, temperature and transport conditions of ocular tissue: Following slaughter, the intact chicken heads were placed into individual plastic compartments within a plastic box in order to minimize any damage to the eyes. The base of each compartment was lined with a paper towel moistened with isotonic saline. The heads were transported to the test facility at ambient temperature.
- Time interval prior to initiating testing: Heads were removed immediately after the chickens had been humanely killed at the source, for use on the same day. The time interval between collection of chicken heads and placing the eyes in the superfusion chamber following enucleation was minimized although all eyes had to fall within the acceptance criteria identified in the test guideline.
- Indication of any existing defects or lesions in ocular tissue samples: no
- Indication of any antibiotics used: no

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 30 µL

Duration of treatment / exposure:

10 sec

Duration of post- treatment incubation (in vitro):

240 min
Reading time points: 30, 75, 120, 180 and 240 min (± 5 min)

Number of animals or in vitro replicates:

3 eyes for test group and positive control; 2 eyes for negative control

Details on study design:

SELECTION AND PREPARATION OF ISOLATED EYES
The integrity of the cornea was measured with a microscope after instillation of sodium fluorescein (2%, w/v). Acceptable values for fluorescein retention and corneal opacity scores are ≤ 0.5. Eyes acceptable for testing were dissected from the skull and pulled from the orbit by holding the nictitating membrane firmly with surgical forceps. The tissue behind the eye was carefully removed with bent, blunt-tipped scissors.
Once the eye was removed from the orbit a portion of the optic nerve remained. Other connective tissue was removed from the eye on an absorbent tray liner. Enucleated eyes were transferred to an appropriate clamp keeping the cornea vertical. They were then transferred to chambers within the superfusion apparatus ensuring the corneas received sufficient isotonic saline from the saline drip. The temperature of the chambers was at 32 ± 1.5 °C.
Once all eyes were placed in the superfusion apparatus, the eyes were examined again for corneal damage with the microscope. Corneal thickness measurements are taken with a depth measuring device on a microscope at the center of each cornea.
Eyes were replaced when: (i) the fluorescein score was > 0.5; (ii) the corneal opacity score was > 0.5; or (iii) there was any additional signs of damage, (iv) the corneal thickness measurements for individual eyes deviated more than 10% from the mean value for all eyes.

EQUILIBRATION AND BASELINE RECORDINGS
After the approval process the eyes were incubated for 45 min for equilibrium purposes. Time zero measurements for corneal thickness and opacity were taken to serve as a baseline. The baseline for the fluorescein measurements were taken at dissection.

REMOVAL OF TEST SUBSTANCE
- Volume and washing procedure after exposure period: The test item was then rinsed from the eye using 20 mL of isotonic saline.

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: Haag-Streit BQ 900 slit-lamp microscope
- Damage to epithelium based on fluorescein retention: Haag-Streit BQ 900 slit-lamp microscope
- Swelling: Corneal thickness was measured with a depth measuring device no. 1 on a slit-lamp microscope.
- Macroscopic morphological damage to the surface: Haag-Streit BQ 900 slit-lamp microscope


SCORING SYSTEM:
- Mean corneal swelling (%) : Percentage corneal swelling was assessed from corneal thickness measurements. The calculation was expressed in the following formula:
(corneal thickness at time (t) - corneal thickness at time =0) / (corneal thickness at time =0) x 100
Mean percentage of corneal swelling for all test eyes was calculated for all the time points. The overall category score was determined from the highest mean score for epithelial swelling as observed at any time point.

- Mean maximum opacity score :
Corneal opacity was calculated with the most densely opacified areas for scoring. The mean value for all test eyes was calculated for all time points. The highest mean score, as observed at any time point was given an overall category for each test item.

Opacity scores:
0 = No opacity
0.5 = Very faint opacity
1 = Scattered or diffuse areas; details of the iris are clearly visible
2 = Easily discernible translucent area; details of the iris are slightly obscured
3 = Severe corneal opacity; no specific details of the iris are visible; size of the pupil barely discernible
4 = Complete corneal opacity; iris invisible

- Mean fluorescein retention score at 30 minutes post-treatment :
The mean fluorescein retention scores for all test eyes are calculated at the 30 minute time interval only. These measurements are used for the overall classification for each test item.

Fluorescein retention scores:
0 = No fluorescein retention
0.5 = Very minor single cell staining
1 = Single cell staining scattered throughout the treated area of the cornea
2 = Focal or confluent dense single cell staining
3 = Confluent large areas of the cornea retaining fluorescein

DECISION CRITERIA:
ICE classification criteria as explained in OECD TG 438 were used.

Irritation parameter:

cornea opacity score

Run / experiment:

240 min

Value:

1

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: ICE Class II

Irritation parameter:

fluorescein retention score

Run / experiment:

30 min

Value:

0.7

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: ICE Class II

Irritation parameter:

percent corneal swelling

Run / experiment:

240 min

Value:

12.44

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: ICE Class II

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: Not specified

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes, No fluorescein retention was noted in both negative control treated eyes.
- Acceptance criteria met for positive control: Yes, Confluent large areas of the cornea retaining fluorescein were noted in all positive control treated eyes.

Table 2: Ocular reactions in observed eyes

	Score	ICE class
Test item
Maximal mean score for corneal opacity	1.0	II
Mean score of fluorescein retention	0.7	II
Maximan mean corneal swelling	12.44%	II
Positive control
Maximal mean score for corneal opacity	4.0	IV
Mean score of fluorescein retention	3.0	IV
Maximan mean corneal swelling	47.57%	IV
Negative control
Maximal mean score for corneal opacity	0.3	I
Mean score of fluorescein retention	0.0	I
Maximan mean corneal swelling	0.00%	I

Interpretation of results:

other: non-corrosive according to OECD 438

Conclusions:

The irritation potential of the test substance was assessed in the ICE assay. After treatment with the neat test substance corneal opacity, fluorescein retention and corneal swelling were all classed as category II. According to OECD 438 the test substance does not have to be classified as corrosive to the eye but no prediction on the eye irritation potential can be made and further testing is required for classification and labeling purposes.

Reference 2

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

04 Jul - 31 Aug 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)

Version / remarks:

adopted: Jul 2015

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Hess. Ministerium für Umwelt, Klimaschutz, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany

Species:

human

Strain:

other: EpiOcular™

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 50 µL

NEGATIVE CONTROL
- Amount applied: 50 µL

POSITIVE CONTROL
- Amount applied: 50 µL

Duration of treatment / exposure:

30 min

Duration of post- treatment incubation (in vitro):

post-exposure immersion: 12 min
post-treatment incubation: 120 min

Number of animals or in vitro replicates:

in duplicates for each treatment and contol group

Details on study design:

- RhCE tissue construct used, including batch number: EpiOcular™ (MatTek Corporation, Bratislava, Slovakia), Lot No.: 23799 (main experiment 1) and 27002 (main experiment 2)
- Tissue viability: The quality of the final product was assessed by undertaking an MTT cell viability test. The determined OD (540 - 570 nm) was 1.688 ± 0.052 and 1.269 ± 0.039 (acceptance criteria: 1.1 - 3.0) for tissues of main experiments 1 and 2, respectively.
- Barrier function: The barrier function was assessed by determination of the exposure time required to reduce tissue viability by 50% (ET-50) upon application of 100 µL of 0.3% Triton X-100. The ET-50 value was determined to be 24.25 min and 24.36 min (acceptance criteria: 12.2 - 37.5 min) for tissues of main experiments 1 and 2, respectively.
- Contamination: The cells used to produce the EpiOcular tissue were screened for the presence of viruses, bacteria, yeast and other fungi.
- Duration and temperature of exposure, post-exposure immersion and post-exposure incubation periods: 30 min exposure (37 °C), 12 min post-exposure immersion (room temperature), 120 min post-exposure (37 °C)
- Indication of controls used for direct MTT-reducers and/or colouring test chemicals: The optical pre-experiment (colour interference pre-experiment) to investigate the test item’s colour change potential in water or isopropanol did not led to a change in colour. Therefore, an additional test with viable tissues without MTT addition was not necessary. Optical evaluation of the MTT-reducing capacity of the test item with MTT-reagent did not show blue colour. Therefore, an additional test with freeze-killed tissues was not necessary.
- Description of the method used to quantify MTT formazan: The absorbance at 570 nm of each well was measured with a plate reader (Versamax Molecular Devices, Ismaning, Germany). No reference wavelength measurement was used.
- Decision criteria/Prediction model: If the test item-treated tissue viability is > 60% relative to the negative control treated tissue viability, the test item is labeled non-irritant. If the test item-treated tissue viability is ≤ 60% relative to negative control treated tissue viability, the test item is labeled irritant.
- Acceptance criteria: The results are acceptable if 1) the negative control OD is > 0.8 and < 2.5, 2) the mean relative viability of the positive control is below 50% of the negative control viability and 3) the difference of viability between the two relating tissues of a single test substance is < 20% in the same run (for positive and negative control tissues and tissues of test substances).

Irritation parameter:

other: % mean relative absorbance of 2 tissues

Run / experiment:

30 min

Value:

61.6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: Experiment 1

Irritation parameter:

other: % mean relative absorbance of 2 tissues

Run / experiment:

30 min

Value:

40.3

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: Experiment 2 (confirmatory experiment)

Other effects / acceptance of results:

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The negative control OD (1.498 and 1.620 for experiment 1 and 1.674 and 1.753 for experiment 2) was in the range of > 0.8 and < 2.5.
- Acceptance criteria met for positive control: The mean relative viability of the positive control is below 50% of the negative control viability (40.1% and 36.5% for experiment 1 and 2, respectively).

The difference of viability between the two relating tissues of a single item is < 20% (values between 1.3% to 4.2% and between 0.6% to 13.0% for experiment 1 and 2, respectively) in the same run (for positive and negative control tissues and tissues of single test items).

Table 2. Experiment 1: Results after treatment for 30 minutes with the test substance and controls

Dose Group	Ab-sorbance Well 1 (Tissue 1/2)	Ab-sorbance Well 2 (Tissue 1/2)	Mean Absor-bance (Tissue 1/2)	Mean Absorbance Tissue 1 and 2 blank corrected	Mean Absorbance of 2 Tissues	Rel. Absorbance [%] Tissue 1 and 2*	Absolute Value of the Difference of the Rel. Absorbances [%] Tissue 1 and 2	Mean Rel. Absorbance [%]**
Blank	0.035	0.035	0.035
Negative Control	1.498	1.582	1.540	1.505	1.533	98.2	3.7	100.0
	1.573	1.620	1.596	1.561		101.8
Positive Control	0.638	0.643	0.641	0.606	0.615	39.5	1.3	40.1
	0.661	0.659	0.660	0.625		40.8
Test Item	0.989	1.034	1.012	0.976	0.944	63.7	4.2	61.6
	0.952	0.941	0.946	0.911		59.4

*  Relative absorbance [rounded values]: 100 x [absorbance of test item or controls / mean absorbance of negative control]

** Mean relative absorbance [rounded values]: 100 x [mean absorbance of test item or controls / mean absorbance of negative control]

Table 2. Experiment 2 (confirmatory experiment): Results after treatment for 30 minutes with the test substance and controls

Dose Group	Ab-sorbance Well 1 (Tissue 1/2)	Ab-sorbance Well 2 (Tissue 1/2)	Mean Absor-bance (Tissue 1/2)	Mean Absorbance Tissue 1 and 2 blank corrected	Mean Absorbance of 2 Tissues	Rel. Absorbance [%] Tissue 1 and 2*	Absolute Value of the Difference of the Rel. Absorbances [%] Tissue 1 and 2	Mean Rel. Absorbance [%]**
Blank	0.038	0.038	0.038
Negative Control	1.674	1.753	1.714	1.676	1.691	99.1	1.8	100.0
	1.735	1.753	1.744	1.706		100.9
Positive Control	0.626	0.693	0.660	0.622	0.617	36.8	0.6	36.5
	0.647	0.652	0.649	0.611		36.2
Test Item	0.610	0.610	0.610	0.572	0.682	33.8	13.0	40.3
	0.827	0.832	0.829	0.792		46.8

* Relative absorbance [rounded values]: 100 x [absorbance of test item or controls / mean absorbance of negative control]

** Mean relative absorbance [rounded values]: 100 x [mean absorbance of test item or controls / mean absorbance of negative control]

Interpretation of results:

other: Eye irritating potential according to OECD 492

Conclusions:

Under the conditions of the conducted test, the test substance is irritating towards human-derived epidermal keratinocytes in the EpiOcular™ model.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin

The skin irritation potential of the test substance was determined by an in vivo skin irritation test in rabbits according to OECD Guideline 404 and in compliance with GLP (1991). Four rabbits were treated each with 0.5 mL of the test substance at concentrations of 1, 5, 10 and 25% diluted in ethanol/diethylphthalate (1:1), undiluted test substance and the vehicle under semi-occlusive dressing for 4 h. Treatment with the undiluted test substance caused skin irritation (mean erythema values over 24, 48 and 72 h: 2.67, 2.67, 3.00, 2.67; mean edema values over 24, 48 and 72 h: 2.67, 1.67, 1.67, 2.33). Signs of inflammation (scaling) were not fully reversible after 21 days. Based on the available data, the test substance is considered to be a skin irritant.

 

Eye

The eye irritation potential of the test substance was determined in an isolated chicken eye test (ICE) according to OECD Guideline 438 and in compliance with GLP (2017). Three isolated chicken eyes per group were exposed to a single application of the undiluted test substance for 10 sec using an application volume of 30 μL. Additional control groups were treated with physiological saline (negative control) or benzalconium chloride (positive control). Treatment with the test substance caused a mean corneal opacity score of 1.0, a mean fluorescein retention score of 0.7 and a mean corneal thickness of 12.44%. All three observation endpoints resulted in ICE class II. In conclusion, the test substance does not have to be classified as corrosive to the eye but no prediction on the eye irritation potential can be made and further testing is required.

 

The eye irritation potential of the test substance was determined by an in vitro eye irritation test using a human cornea model according to OECD Guideline 492 and in compliance with GLP (2018). 50 µL of the neat test substance, negative (deionized water) and positive control (methyl acetate), respectively, were applied to each of duplicate EpiOcular™ tissues for 30 min. In a first experiment, a borderline result (61.6% tissue viability; threshold for irritation: 60%) was observed following incubation with the test substance. Therefore, a second experiment was performed indicating a clear irritating effect (40.3% tissue viability) of the test substance.

Even if the results were inconsistent between experiment 1 and 2, it was decided not to carry out an additional third experiment as the results of the second experiment were significantly lower than the threshold of 60% and in addition they were close to the results of the positive control.

In conclusion, based on the available data, the test substance is considered to be irritating to the eye.

Justification for classification or non-classification

The available data on skin irritation meet the criteria for classification as Skin Irrit. Cat. 2 (H315) according to Regulation (EC) 1272/2008.

The available data on eye irritation meet the criteria for classification as Eye Irrit. Cat. 2 (H319) according to Regulation (EC) 1272/2008.