Fatty acids, C16-18 (even numbered), reaction products with tetraethylenepentamine, acetates (salts)

Endpoint summary

Description of key information

Skin irritation:
- not irritating in rabbits (OECD guideline 404 / EU method B.4, GLP), applied to the intact skin for 4 hours; exposure to the test substance resulted in very slight or well defined erythema (score 1 - 2) and very slight or slight edema (score 1 - 2) in the treated skin areas; fully reversible within 10 d max.

Eye irritation:
- not severely irritating/not corrosive (OECD 437, BCOP, GLP), 10% suspension, mean in vitro irritation score = 0.85
- irritating in rabbits (OECD guideline 404 / EU method B.4, GLP), based on irreversible effects in a study conducted with the target substance and read-across approach to source substances

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:

25-JAN-2005 - 15-FEB-2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)

Version / remarks:

(29 April 2004)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

(24 April 2002)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

- Name of test material: Fatty acids, C16-18, reaction products with tetraethylenepentamine, acetates (salts)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories France, F-69592 L’Arbresle
- Age at study initiation: 42 weeks (male), 11 weeks (females)
- Housing: Individually in stainless steel cages
- Diet (e.g. ad libitum): Pelleted standard Provimi Kliba 3418 rabbit maintenance diet ad libitum (batch no. 68/04) provided by Provimi Kliba
AG, CH-4303 Kaiseraugst
- Water (e.g. ad libitum): Community tap water from Fillinsdorf, ad libitum.
- Acclimation period: 6-7 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17-23 °C
- Humidity (%): 30-70%
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

unchanged (no vehicle)

Remarks:

moistenes with 0.5 mL purified water

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 g/animal

Duration of treatment / exposure:

4 h

Observation period:

14 d

Number of animals:

3

Details on study design:

TEST SITE
- Area of exposure: 4 cm x 4 cm on the flank
The patch was covered with a semi-occlusive. dressing. The dressing was wrapped around the abdomen and anchored with tape.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): lukewarm tap water
- Time after start of exposure: 4 h

OBSERVATION TIME POINTS
1, 24, 48 and 72 hours, 7, 10 and 14 days

SCORING SYSTEM:
- Method of calculation: Draize scale

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.67

Max. score:

4

Reversibility:

fully reversible within: 7 d

Remarks on result:

other: scaling was observed from the 72 h observation, persisting up to day 10

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1.33

Max. score:

4

Reversibility:

fully reversible within: 10 d

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.67

Max. score:

4

Reversibility:

fully reversible within: 72 h

Irritation parameter:

edema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.67

Max. score:

4

Reversibility:

fully reversible within: 72 h

Remarks on result:

other: scaling was observed from the 72 h observation, persisting up to day 10

Irritation parameter:

edema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 7 d

Irritation parameter:

edema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

 

Score at time point	3 min application		1 h application
	Erythema	Edema	Erythema	Edema
0 h	0	0	1	0
60 min	0	0	1	0
24 h	0	0	1	0
48 h	0	0	1	0
72 h	0	0	1	0
7 d	0	0	0	0
10 d	0	0	0	0
14 d	0	0	0	0

 

Irritant/corrosive response data for each animal at each observation time (4 h treatment):

Score at time point	Erythema	Edema	Scaling
60 min	2/2/2	2/2/2	0/0/0
24 h	2/2/1	1/1/0	0/0/0
48 h	2/1/1	1/1/0	0/0/0
72 h	1/1/0	0/1/0	x/0/0
7 d	0/1/0	0/0/0	x/0/0
10 d	0/0/0	0/0/0	x/0/0
14 d	0/0/0	0/0/0	0/0/0
Average 24, 48, 72 h	1.67/1.33/0.67	0.67/1/0	Not calculated

Interpretation of results:

GHS criteria not met

Conclusions:

In this study C16-18FA-TEPA-compound was not irritating to the skin.

Executive summary:

In a primary dermal irritation study according to OECD guideline 404 (24 April 2002) and EU method B.4 (29 April 2004) 3 young adult New Zealand White rabbits were dermally exposed to 0.5 mg of C16-18FA-TEPA-compound (90% a.i.) for 4 hours. The substance was moistened with 0.5 mL purified water. Animals then were observed for 14 days.  Irritation was scored by the method of Draize.

Very slight to well-defined erythema were observed from the 1- to the 48-hour reading in all animals and persisted as very slight erythema up to the 72-hour or 7-day examination in two animals, respectively. Slight edema was observed in all animals one hour after treatment and very slight edema persisted up to the 48- or 72-hour reading in two animals, respectively. Scaling was present at the application site of one animal from the 72-hour to the 10-day examination.

Mean erythema scores from observations at 24, 48 and 72 h after patch removal were 1.67/1.33/0.67; mean edema scores were 0.67/1/0.

In this study C16-18FA-TEPA-compound was not irritating to the skin.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that source and target substances have similar toxicological and ecotoxicological properties because
• they are manufactured from similar precursors under similar conditions
• they share structural similarities and common functional groups
• the analytical descriptors show comparable results
• the metabolism pathway leads to comparable products (amine backbone and long chain fatty acids) and non-common products predicted to have no toxicological effects (long chain fatty acids).

This read-across hypothesis corresponds to scenario 2 - different compounds have qualitatively and quantitatively the same type of effects - of the read-across assessment framework i.e. properties of the target substance C1618FA-TEPA-compound are predicted to be similar to those of the source substances Partially unsaturated IQAC, DMS quaternised and oleic acid based IQAC, DMS quaternised.

Based on the available experimental data, the read-across strategy is supported by close structural analogy as well as similar toxicological profiles.

Therefore, read-across from the existing sub-chronic toxicity studies and pre-natal developmental toxicity as well as ecotoxicological studies on the source substance is considered as an appropriate adaptation to the standard information requirements of the REACH Regulation for the target substance, in accordance with the provisions of Annex XI, 1.5 of the REACH Regulation.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Please refer to detailed Justification for read-across attached to Iuclid section 13

3. ANALOGUE APPROACH JUSTIFICATION
Please refer to detailed Justification for read-across attached to Iuclid section 13

4. DATA MATRIX
Please refer to detailed Justification for read-across attached to Iuclid section 13

Reason / purpose for cross-reference:

read-across: supporting information

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Species:

rabbit

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

>= 0 - <= 1.67

Max. score:

4

Reversibility:

fully reversible

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

>= 1.33 - <= 2

Max. score:

3

Reversibility:

fully reversible

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

>= 1 - <= 2.33

Max. score:

4

Reversibility:

fully reversible

Interpretation of results:

Category 2 (irritating to eyes) based on GHS criteria

Reference 2

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20-Feb-2013 - 20-Feb-2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

Version / remarks:

30 May 2008

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

adopted 24th April 2002

GLP compliance:

yes

Specific details on test material used for the study:

- Name of test material: Fatty acids, C16-18, reaction products with tetraethylenepentamine, acetates (salts)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Charles River Germany, Sandhoferweg 7, 97633 Sulzfeld / Germany
- Age at study initiation: 13 weeks
- weight at study initiation: 2341 - 3038 g
- Housing: individually
- Diet (e.g. ad libitum): Pelleted standard Teklad Global High Fiber Rabbit Diet 2031C, ad libitum
- Water (e.g. ad libitum): Community tap water, ad libitum
- Acclimation period: no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

unchanged (no vehicle)

Controls:

other: the untreated right eye remained untreated and served as the reference control

Duration of treatment / exposure:

1 h (eyes were rinsed with 0.9% NaCl 1 h after start of exposure)

Observation period (in vivo):

18 d

Number of animals or in vitro replicates:

3

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The treated eyes were rinsed with NaCl 0.9% 1 hour after instillation.

SCORING SYSTEM: Draize scale

TOOL USED TO ASSESS SCORE: hand-slit lamp / biomicroscope / fluorescein

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Max. score:

3

Reversibility:

fully reversible within: 18 d

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

2.67

Max. score:

4

Reversibility:

fully reversible within: 18 d

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

2

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

2

Reversibility:

not reversible

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

3

Reversibility:

not reversible

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1

Max. score:

2

Reversibility:

not reversible

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1

Max. score:

3

Reversibility:

not reversible

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

3.33

Max. score:

4

Reversibility:

not reversible

Irritant / corrosive response data:

The instillation of Fatty acids, C16-C18 (even numbered), reaction products with
tetraethylenepentamine, acetates into the eye resulted severe ocular changes, such as reddening of the conjunctivae and sclera, ocular discharge, chemosis, corneal opacity and sluggish iris light reflex. Eye effects persisted until 17 days after treatment in animal no. 1 but were irreversible in animal nos. 2 and 3. At several time points, the sclera of animal nos. 2 and 3 could not be assessed due to swollen conjunctivae.
Fluorescein staining was positive in animal no. 1 48 and 72 hours as well as 7, 10 and 14 days after test item instillation. For animal nos. 2 and 3, positive fluorescein staining was observed at all observation time points after instillation.
The mean score was calculated separately for each animal across three scoring times (24, 48 and 72 hours after instillation) for corneal opacity, iris light reflex, redness and chemosis of the conjunctivae, respectively. The individual mean scores for corneal opacity and iris light reflex were 0.00, 2.00, 2.00 and 0.00, 1.00, 1.00, respectively. The individual mean scores for the conjunctivae were 1.00, 1.00 and 1.00 for reddening and 2.67, 3.00 and 3.33 for chemosis, respectively.

Other effects:

No intercurrent deaths occurred during the course of the study.

No clinical signs were recorded throughout the entire observation period.

The body weight of the animals was within the range commonly recorded for this strain and age.

Although the treated eyes of all animals were thoroughly flushed with 0.9% NaCl one hour after test item instillation, some test item remnants remained in the eyes of animal nos. 2 and 3. These test item remnants could neither be removed by flushing the eyes with NaCl 0.9%, nor mechanically using a paper tissue and fingers and stuck to the conjunctivae until 72 hours after instillation in animal no. 3 and until 10 days after instillation in animal no. 2.

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

In this study, C1618FA-TEPA-compound caused irreversible effects to the eye (Category 1).

Executive summary:

In a primary eye irritation study according to OECD guideline 405 (adopted 24th April 2002) and EU method B.5 (30 May 2008) 0.1 g of C1618FA-TEPA-compound (100% a.i.) was instilled into the conjunctival sac of the left eye of each of three young adult New Zealand White rabbits. Scoring of irritation effects was performed approximately 1, 24, 48 and 72 hours, as well as 7, 10, 14, 17 and 21 days after test item instillation. Corneal opacity was additionally assessed by fluorescein staining. At first, only one animal was treated. After 1 h, the eye was rinsed with 0.9% NaCl. The animal was observed for 18 d. Irritation was scored by the method of Draize.

The instillation of the test item into the eye resulted in severe ocular changes, such as reddening of the conjunctivae and sclera, ocular discharge, chemosis, corneal opacity and sluggish iris light reflex. Eye effects persisted until 17 days after treatment in animal no. 1 but were irreversible in
animal nos. 2 and 3. At several time points, the sclera of animal nos. 2 and 3 could not be assessed due to swollen conjunctivae.
Fluorescein staining was positive in animal no. 1 48 and 72 hours as well as 7, 10 and 14 days after test item instillation. For animal nos. 2 and 3, positive fluorescein staining was observed at all observation time points after instillation.

The mean score was calculated separately for each animal across three scoring times (24, 48 and 72 hours after instillation) for corneal opacity, iris light reflex, redness and chemosis of the conjunctivae, respectively. The individual mean scores for corneal opacity and iris light reflex were 0.00, 2.00, 2.00 and 0.00, 1.00, 1.00, respectively. The individual mean scores for the conjunctivae were 1.00, 1.00 and 1.00 for reddening and 2.67, 3.00 and 3.33 for chemosis, respectively.

Although the treated eyes of all animals were thoroughly flushed with 0.9% NaCl one hour after test item instillation, some test item remnants remained in the eyes of animal nos. 2 and 3. These test item remnants could neither be removed by flushing the eyes with NaCl 0.9%, nor
mechanically using a paper tissue and fingers and stuck to the conjunctivae until 72 hours after instillation in animal no. 3 and until 10 days after instillation in animal no. 2.

In this study, C1618FA-TEPA-compound caused irreversible effects to the eye (Category 1).

Reference 3

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

January 25, 2013 - January 25, 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

December 2010

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

September 2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

- Name of test material: Fatty acids, C16-18, reaction products with tetraethylenepentamine, acetates (salts)

Species:

cattle

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Source: Freshly isolated bovine cornea; Schlachthof Bensheim, 64625 Bensheim, Germany
- Characteristics of donor animals (e.g. age, sex, weight): at least 9 month old donor cattle
The isolated eyes were transported to the laboratory in Hank’s BSS supplemented with streptomycin / penicillin at ambient temperature. The corneae were isolated on the same day after delivery of the eyes and used directly for the BCOP test

Vehicle:

physiological saline

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

0.75 mL as 10% (w/v) suspension of the test item in saline

Duration of treatment / exposure:

The corneas were exposed to the test item solution for 10 minutes.

Duration of post- treatment incubation (in vitro):

2 h

Number of animals or in vitro replicates:

3 corneas per group (test item, negative control, positive control)

Details on study design:

NUMBER OF REPLICATES: 3

NEGATIVE CONTROL USED: 0.9% (w/v) NaCl in deionised water

POSITIVE CONTROL USED: 2-Ethoxyethanol

APPLICATION DOSE AND EXPOSURE TIME

TREATMENT METHOD: open chamber

POST-INCUBATION PERIOD: 2 h

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: OP_KiT opacitometer
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of UV/VIS spectrophotometry (OD490)

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA: In vitro Irritation Score = opacity value + (15 x OD490 value)
IVIS>/= 55.1: Corrosive / severe irritant CLP/EPA/GHS (Cat 1)

VALIDITY CRITERIA: The test was acceptable if the in vitro irritation score of the positive control was ≥ 30 and the in vitro irritation score of the negative control was ≤ 3.

Irritation parameter:

in vitro irritation score

Value:

0.85

Negative controls validity:

valid

Positive controls validity:

valid

Results after 10 Minutes Incubation Time

Test Group	Opacity value = Difference (t130-t0) of Opacity		Permeability at 490 nm (OD490)		In vitro Score	Mean in vitro Irritation Score	Proposed in vitro Irritation Scale
		Mean		Mean
Negative Control	1	0.67	0.048	0.048	1.72	1.38	Non corrosive / non severe irritant
	0		0.047		0.71
	1		0.048		1.72
Positive Control	51.33*		0.191*		54.20	59.15	Corrosive / severe irritant
	60.33*		0.285*		64.61
	54.33*		0.286*		58.63
10% (w/v) suspension of the test item in saline	0.33*		0.031*		0.80	0.85	Non corrosive / non severe irritant
	0.33*		0.036*		0.88
	0.33*		0.035*		0.86

*corrected values

Interpretation of results:

other: not corrosive

Conclusions:

In conclusion, according to the current study and under the experimental conditions reported, the 10% (w/v) suspension of C16-18FA-TEPA-compound in saline is not corrosive to the eye.

Executive summary:

This in vitro study was performed to assess the corneal irritation and damage potential of C16-18FA-TEPA-compound by means of the BCOP assay using fresh bovine corneas. The test item was tested as 10% (w/v) suspension in saline. The positive control 2-Ethoxyethanol was tested neat. Saline was used as negative control item.

Prior to the test the test item was crushed in a mortar with a pistil to improve its consistency. Following, a 10% (w/v) suspension of the test item in saline was prepared. Although using ultra sonic technique, the test item could not be suspended homogeneously. Therefore, 0.75 mL of the vehicle/test item mixture were applied onto the corneas taking care, that the test item was spread evenly across the corneas, and that the test item got in contact with the corneas.

After a first opacity measurement of the fresh bovine corneas (t0), each 0.75 mL of the 10% (w/v) suspension of the test item in saline, of the positive control, and of the negative control were applied to the corneas and incubated for 10 minutes at 32 ± 1°C. After the incubation phase the test item suspension, the positive, and the negative controls were each rinsed from the corneas. Further, the corneas were incubated for another 120 minutes at 32 ± 1°C in complete medium, and opacity was measured a second time (t130).

After the opacity measurements permeability of the corneas was determined by measuring spectrophotometrically the transfer of fluorescein after incubation in a horizontal position for 90 minutes at 32 ± 1°C.

With the negative control (saline) neither an increase of opacity nor permeability of the corneas could be observed (mean in vitro irritation score 1.38).

The positive control (2-Ethoxyethanol) showed clear opacity and distinctive permeability of the corneas (mean in vitro irritation score 59.15) corresponding to a classification ascorrosive /severe irritant to the eye (CLP/EPA/GHS (Cat 1)).

Relative to the negative control, the 10% (w/v) suspension of C16-18FA-TEPA-compound in saline did not induce any increase of the corneal opacity or permeability. The calculated mean in vitro irritation score was 0.85. According to OECD 437 the 10% (w/v) suspension of the test item in saline is classified as not corrosive / not severe irritant to the eye.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation

In a primary dermal irritation study according to OECD guideline 404 (24 April 2002) and EU method B.4 (29 April 2004) 3 young adult New Zealand White rabbits were dermally exposed to 0.5 mg of C1618FA-TEPA-compound (90% a.i.) for 4 hours. The substance was moistened with 0.5 mL purified water. Animals then were observed for 14 days.  Irritation was scored by the method of Draize.

Very slight to well-defined erythema were observed from the 1- to the 48-hour reading in all animals and persisted as very slight erythema up to the 72-hour or 7-day examination in two animals, respectively. Slight edema was observed in all animals one hour after treatment and very slight edema persisted up to the 48- or 72-hour reading in two animals, respectively. Scaling was present at the application site of one animal from the 72-hour to the 10-day examination.

Mean erythema scores from observations at 24, 48 and 72 h after patch removal were 1.67/1.33/0.67; mean edema scores were 0.67/1/0.

In this study C1618FA-TEPA-compound was not irritating to the skin.

 

Eye irritation

The eye irritation potential of C1618FA-TEPA-compound was assessed in a weight of evidence approach using an in vitro test and a not yet finalized in vivo study conducted with the target substance as well as supporting information from structurally closely related substances (partially unsaturated IQAC, DMS quaternised; oleic-acid based IQAC, DMS quaternised). A justification for read-across isattached to Iuclid section 13.

 

An his in vitro study was performed to assess the corneal irritation and damage potential of C1618FA-TEPA-compound by means of the BCOP assay using fresh bovine corneas. The test item was tested as 10% (w/v) suspension in saline. The positive control 2-Ethoxyethanol was tested neat. Saline was used as negative control item.

Prior to the test the test item was crushed in a mortar with a pistil to improve its consistency. Following, a 10% (w/v) suspension of the test item in saline was prepared. Although using ultra sonic technique, the test item could not be suspended homogeneously. Therefore, 0.75 mL of the vehicle/test item mixture were applied onto the corneas taking care, that the test item was spread evenly across the corneas, and that the test item got in contact with the corneas.

After a first opacity measurement of the fresh bovine corneas (t0), each 0.75 mL of the 10% (w/v) suspension of the test item in saline, of the positive control, and of the negative control were applied to the corneas and incubated for 10 minutes at 32 ± 1°C. After the incubation phase the test item suspension, the positive, and the negative controls were each rinsed from the corneas. Further, the corneas were incubated for another 120 minutes at 32 ± 1°C in complete medium, and opacity was measured a second time (t130).

After the opacity measurements permeability of the corneas was determined by measuring spectrophotometrically the transfer of fluorescein after incubation in a horizontal position for 90 minutes at 32 ± 1°C.With the negative control (saline) neither an increase of opacity nor permeability of the corneas could be observed (meanin vitroirritation score 1.38). The positive control (2-Ethoxyethanol) showed clear opacity and distinctive permeability of the corneas (meanin vitroirritation score 59.15) corresponding to a classification ascorrosive /severe irritant to the eye (CLP/EPA/GHS (Cat 1)).

Relative to the negative control, the 10% (w/v) suspension of C1618FA-TEPA-compound in saline did not induce any increase of the corneal opacity or permeability. The calculated mean in vitro irritation score was 0.85. According to OECD 437 the 10% (w/v) suspension of the test item in saline is classified as not corrosive / not severe irritant to the eye.

 

In a primary eye irritation study according to OECD guideline 405, 0.1 g of C1618FA-TEPA-compound (100% a.i.) was instilled into the conjunctival sac of the left eye of each of three young adult New Zealand White rabbits. Scoring of irritation effects was performed approximately 1, 24, 48 and 72 hours, as well as 7, 10, 14, 17 and 21 days after test item instillation. Corneal opacity was additionally assessed by fluorescein staining. At first, only one animal was treated. After 1 h, the eye was rinsed with 0.9% NaCl. The animal was observed for 18 d. Irritation was scored by the method of Draize.

The instillation of the test item into the eye resulted in severe ocular changes, such as reddening of the conjunctivae and sclera, ocular discharge, chemosis, corneal opacity and sluggish iris light reflex. Eye effects persisted until 17 days after treatment in animal no. 1 but were irreversible in
animal nos. 2 and 3. At several time points, the sclera of animal nos. 2 and 3 could not be assessed due to swollen conjunctivae.
Fluorescein staining was positive in animal no. 1 48 and 72 hours as well as 7, 10 and 14 days after test item instillation. For animal nos. 2 and 3, positive fluorescein staining was observed at all observation time points after instillation.

The mean score was calculated separately for each animal across three scoring times (24, 48 and 72 hours after instillation) for corneal opacity, iris light reflex, redness and chemosis of the conjunctivae, respectively. The individual mean scores for corneal opacity and iris light reflex were 0.00, 2.00, 2.00 and 0.00, 1.00, 1.00, respectively. The individual mean scores for the conjunctivae were 1.00, 1.00 and 1.00 for reddening and 2.67, 3.00 and 3.33 for chemosis, respectively.

Although the treated eyes of all animals were thoroughly flushed with 0.9% NaCl one hour after test item instillation, some test item remnants remained in the eyes of animal nos. 2 and 3. These test item remnants could neither be removed by flushing the eyes with NaCl 0.9%, nor
mechanically using a paper tissue and fingers and stuck to the conjunctivae until 72 hours after instillation in animal no. 3 and until 10 days after instillation in animal no. 2.

In this study, C1618FA-TEPA-compound caused irreversible effects to the eye (Category 1).

 

 

In a primary eye irritation study according to OECD Guideline 405 (2002), 0.1 mL of partially unsaturated IQAC, DMS quaternised (75 % in propylene-glycol) was instilled into the conjunctival sac of three New Zealand White rabbits. The eyes were not rinsed after substance application. Animals then were observed for 7 days. Irritation was scored by the method of Draize. At 24 hours post application the eyes of all animals were further examined with fluorescein for cornea damage. All three animals showed moderate ocular reaction 1 h after application of the test substance (two scores for conjunctivae redness and chemosis). 24 h after application moderate ocular reactions were still observed in all three animals. Cornea opacity or iris irritation was not observed in any of the three animals. Slight irritation of the conjunctivae persisted until 72 h after treatment in all animals but 7 days after application all three animals were without any sign of ocular irritation.

 

In a primary eye irritation study according to US Guideline EPA OPPTS 870.2400 (1998), OECD Guideline 405 (1987), EU Guideline B.5 (1992) and Japanese Guideline Nousan No. 6283 (1999), revised Nousan 8628 (2000), 0.1 mL ofoleic-acid based IQAC, DMS quaternised (according to sponsors information: 80 % of CAS No. 72749-55-4, 15 % of CAS No. 9004-96-0 (Oleic acid, ethoxylated) and 5 % of CAS No. 25322-68-3 (Poly(oxy-1,2-ethanediyl),α-hydro-ω-hydroxy- Ethane-1,2-diol, ethoxylated)) was instilled into the conjunctival sac of three White New Zealand rabbits. The eyes were not rinsed after substance application. Animals then were observed for 21 days. Irritation was scored by the method of Draize. Positive corneal, iridal and conjunctival findings were noted for all animals. Corneal neovascularization was noted for two animals on study days 7 through 14 or 17. Iridal irritation subsided by study day 7, and corneal and conjunctival irritation completely subsided by study termination (study day 21). The test substance induced a maximum mean total score of 50.3 (maximum possible mean total score of 110) at 48 hours post-instillation. Mean scores following grading at 24, 48 and 72 hours after instillation of the test material for corneal opacity were >/= 1but < 3, for iritis >/= 1 but < 1.5 and for conjunctival redness and oedema >/= 2. All effects were completely reversible in all animals within 21 days.

 

Based on the overall data on eye irritation (not corrosive in the BCOP assay, irreversible effects in the in vivo eye irritation test), C1618FA-TEPA-compound is considered to be irritating to the eyes. This is supported by eye irritation data on closely related source substances. The higher unsaturated source substanceoleic-acid based IQAC, DMS quaternised can be regarded as an overall worst case as higher degrees of unsaturation usually correlate with higher irritating potential. Thus, by classifying the target substance C1618FA-TEPA-compound as eye irritant (Category 2) a sufficient degree of conservatism is applied.

 

Respiratory irritation

No data on the respiratory irritation of C1618FA-TEPA-compound are available.

 

There are no data gaps for the endpoint irritation/corrosion. No human information is available for this endpoint. However, there is no reason to believe that these results would not be applicable to humans.

 

Justification for classification or non-classification

Skin irritation

Based on the available data, C1618FA-TEPA-compound does not need to be classified for skin irritation according to the criteria of regulation (EC) 1272/2008.

 

 

Eye irritation

Based on reliable, adequate and relevant data, C1618FA-TEPA-compound is classified as irritating to the eyes (Category 2) according to CLP, EU GHS (Regulation (EC) No 1272/2008) and is assigned the hazard statement H319 and the signal word “Warning”.