Decanoic acid, mixed esters with heptanoic acid, isovaleric acid, octanoic acid and pentaerythritol

Endpoint summary

Description of key information

Skin Corrosion/Irritation

The available information from the read-across substances with in vivo and in vitro data, indicate that the substance is not expected to be corrosive or irritating to skin.

Eye Irritation

Read across to the read-across substances in vivo data suggest that the substance would not cause damage or irritate the eyes.

Bovine Corneal Opacity and Permeability (BCOP) Test

Hatcol 1570 showed no effects on the cornea of the bovine eye. The calculated IVIS (in vitro irritancy score) is 0.41.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

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

Remarks:

1 substance available for read-across

Adequacy of study:

key study

Justification for type of information:

See the full attached justification in section 13 for details.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Irritation / corrosion parameter:

% tissue viability

Value:

96.4

Vehicle controls validity:

not applicable

Negative controls validity:

not applicable

Positive controls validity:

valid

Remarks on result:

other: OECD 431

Remarks:

CAS 68130-53-0

Irritation / corrosion parameter:

% tissue viability

Value:

96.4

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: Basis: mean. Time point: 60 minutes. Max. score: 100.0. Remarks: Viability score are reported as %. The lower the score the more harmful a substance is expected to be.

Remarks:

OECD 439 (CAS 68130-53-0)

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minutes exposure

Value:

85.2

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: CAS 68130-53-0

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minute exposure

Value:

100

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: CAS 68130-53-0

Interpretation of results:

GHS criteria not met

Conclusions:

The substance, CAS 68130-51-8; EC 268-594-6, is analogous to the substances to be read across to, in terms of basic form, and the degree of substitution of functional groups is not considered to effect the proposed read across for in vitro skin irritation. The substance is not considered to be corrosive or irritating on the basis of read across.

Reference 2

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with restrictions. Limited documentation but relevant data given.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

Adopted 24 April 2002

Deviations:

yes

Remarks:

limited documentation, limited information on test substance given, 2 h exposure only

GLP compliance:

not specified

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight at study initiation: mean: 3.7 kg

Type of coverage:

occlusive

Preparation of test site:

not specified

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.5 mL
- Concentration: 100%

Duration of treatment / exposure:

2 h

Observation period:

72 h
Reading time points: 1, 24, 48 and 72 h

Number of animals:

5 males

Details on study design:

TEST SITE
- Area of exposure: 9 cm²
- Type of wrap if used: the treated skin was covered with occlusive tape.

REMOVAL OF TEST SUBSTANCE
- Washing: Residual test material was removed with water
- Time after start of exposure: 2 h

SCORING SYSTEM: Draize scoring system

Irritation parameter:

erythema score

Basis:

mean

Remarks:

out of all 5 animals

Time point:

24/48/72 h

Score:

0.24

Max. score:

4

Reversibility:

fully reversible within: 72 h

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Remarks:

out of all 5 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

At tape removal and after 1 h, no effects on the skin were observed. After 24 h, 3 animals showed erythema, which persisted in 1 animal for 48 h. 72 h after exposure, all animals were free of erythema.
Edema were not observed at all.

Interpretation of results:

GHS criteria not met

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Reference 3

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03 Nov - 06 Nov 1987

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP-guideline study.

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

September 1984

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: The Broekman Institute, Someren, The Netherlands
- Age at study initiation: 20 - 21 weeks
- Weight at study initiation: 3.1 kg ± 202.53 g
- Housing: individually in plastic cages with perforated floors
- Diet: 100 g / day, LK-01, Hope Farms, Woerden, The Netherlands
- Water: tap-water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 21
- Humidity (%): 55 -65
- Air changes: air conditioned room
- Photoperiod (hrs dark / hrs light): 12 / 12

Type of coverage:

semiocclusive

Preparation of test site:

other: clipping

Vehicle:

unchanged (no vehicle)

Controls:

other: untreated site of the same animal served as control

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.5 mL
- Concentration: 100 %

Duration of treatment / exposure:

4 h

Observation period:

72 h
Reading time points: 50 min, 24, 48, 72 h

Number of animals:

3 females

Details on study design:

TEST SITE
- Area of exposure: 6 cm²
- Type of wrap: gauze patch attached with a drop of petrolatum to aluminium foil and mounted on permeable tape, held in place with flexible bandage.

REMOVAL OF TEST SUBSTANCE
- Washing: remaining test substance was removed, using a dry tissue and subsequently a tissue moistened with tap-water.
- Time after start of exposure: 4 h

SCORING SYSTEM: Draize scoring system

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.66

Max. score:

4

Reversibility:

fully reversible within: 72 h

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritation parameter:

edema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritation parameter:

edema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritation parameter:

edema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritant / corrosive response data:

Very slight erythema and edema were observed in all animals 24 h after treatment which were fully reversible latest within 72 h.

Other effects:

No signs of systemic toxicity were observed in any of the animals.

Interpretation of results:

GHS criteria not met

Conclusions:

CLP: not classified
DSD: not classified

Reference 4

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13 Nov - 16 Nov 1990

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP guideline study.

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

adopted in 1981

Deviations:

yes

Remarks:

Lack of details on test substance

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Elevage Cunicole du Val de Selle, France
- Weight at study initiation: 2.3 kg
- Diet: ad libitum; Rabbit certified pellet diet (Rabbit sustenance ref. 112 C), ad libitum
- Water: tap water, ad libitum
- Acclimation period: 5 days
-other: the animals received a preventive coccidiosis treatment during acclimation (Mucoxid, 137.5 mg/kg bw/day), via drinking water

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17-23
- Humidity (%): 30-70
- Photoperiod (hrs dark / hrs light): 12 / 12

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

unchanged (no vehicle)

Controls:

other: not required, untreated sites of the same animal served as control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 mL

Duration of treatment / exposure:

4 h

Observation period:

72 h
Reading time points: 1, 24, 48 and 72 h

Number of animals:

3

Details on study design:

TEST SITE
- Area of exposure: 6 cm², flanks were clipped a day before treatment
- Type of wrap if used: adhesive hypoallergic aerated semi-occlusive dressing (Laboratoires de Pansements et d´Hygiène, France)

REMOVAL OF TEST SUBSTANCE
- Washing (if done): No

SCORING SYSTEM: According to Draize

Irritation parameter:

erythema score

Basis:

mean

Remarks:

of all three animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Remarks:

of all three animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

No cutaneous reactions were observed in all the animals.

Erythema score

Animal Number	1 h	24 h	48 h	72 h
1	0	0	0	0
2	0	0	0	0
3	0	0	0	0

 

Edema Score

Animal Number	1 h	24 h	48 h	72 h
1	0	0	0	0
2	0	0	0	0
3	0	0	0	0

 

Interpretation of results:

GHS criteria not met

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Reference 5

Endpoint:

skin corrosion: in vitro / ex vivo

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03 April 2014 to 02 July 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed in accordance with OECD test guidelines in compliance with GLP.

Qualifier:

according to guideline

Guideline:

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

Qualifier:

according to guideline

Guideline:

other: EPA 40 CFR 158.340

GLP compliance:

yes

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: Not specified

Source strain:

not specified

Details on animal used as source of test system:

EpiDerm™ tissues, Lot 19981 Kit D, were received from MatTek Corporation (Ashland, MA) on 08 Apr 2014 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.

Justification for test system used:

In accordance with the test guidelines.

Vehicle:

unchanged (no vehicle)

Details on test system:

Test Article Reduction of MTT: 100 µI of the test article were mixed with 1 ml of MTT 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 MTT. Since tissue Viability is based on MTT 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 MTT and the assay continued as per the protocol.

Mesh Compatibility: Pre-cut nylon meshes supplied with the tissues were placed on a slide and 30 µI of the test article or tissue culture water were 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 µI of the test article were applied to the top of each EpiDerm™ tissue. A nylon mesh was then placed on top to facilitate even distribution of the test article. The test article remained in contact with the EpiDerm™ tissue for 3 and 60 minutes. A Negative Control (TCH2O) and a Positive Control (KOH) were tested at 3 and 60 minutes. A nylon mesh was placed on top of each liquid control to facilitate even distribution of the material. Each treatment with test article or control was conducted in duplicate.
All tissues were dosed at ambient temperature. Tissues treated with the test article and controls for 3 minutes were maintained at ambient temperature until analyzed for Tissue Viability (MTT Reduction). Tissues to be treated for 60 minutes were dosed and then returned to the Incubator (37°C ± 1°C, 5% ± 1% CO2) for the remainder of the 60-minute exposure period

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 MTT solution (1 mg/ml MTT in DMEM). The tissues were then returned to the Incubator for a three-hour MTT incubation period. Following the MTT 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 MTT formazan was measured In triplicate at 540 nm using a microplate reader (µQuant Plate Reader, BioTek Instruments, Winooski, VT).

Analysis of Data: The mean absorbance value for each time point was calculated from the optical density (OD) of the duplicate tissues and expressed as percent viability for each sample using the following formula:
% viability = 100 X (OD sample/OD Negative Control)

Quality Controls: The Negative Controls meets the acceptance criterion if the mean OD of the two tissues at each time point is greater than or equal to 0.8 and is less than or equal to 2.8 (0.8 ≤ OD ≤ 2.8).
The Positive Control meets the acceptance criterion If the mean relative tissue viability at the 60 minute time point is less than 15% < 15%).
Inter-tissue viability meets the acceptance criterion if the difference between two identically treated tissues is no greater than 30%. Re-testing of the chemical is recommended if the resulting viability is near to a classification cut-off. Note: If necessary, the percent difference will be calculated between the mean of the two tissues and one of the tissues. If this difference is greater than 15%, then rejection should be considered.

Interpretation of Results: Skin Corrosion is defined as the production of irreversible tissue damage in skin following the application of test material. The percent viability calculated was used to determine corrosivity potential in the following manner:
Mean Viability Mean Viability
(3 min.) (60 min.) Predicted Corrosivity
<50% Not Applicable Corrosive
≥50% <15% Corrosive
≥50% ≥15% Non-Corrosive

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

50 µI of the test article were applied to the top of each EpiDerm™ tissue.

Duration of treatment / exposure:

3 and 60 minutes

Duration of post-treatment incubation (if applicable):

Not specified

Number of replicates:

Tests were repeated in triplicate.

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minute exposure

Value:

85.2

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minute exposure

Value:

100

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

EXPERIMENTAL DATA

Test Article:		H2925 (CAS No. 68130-53-0), Lot# 2013090401
Dose:		50µl
Conc:		Neat
TIME (min)	OD 1	OD 2	OD 3	MEAN (OD)	SD	VIABILITY %	ERROR %
3.0	1.824	1.833	1.837	1.831	0.007	89.1	0.3
	1.670	1.679	1.671	1.673	0.005	81.4	0.2
Neg control				2.056		100.0
60.0	1.725	1.721	1.717	1.721	0.004	89.4	0.2
	2.112	2.153	2.135	2.133	0.021	110.9	1.1
Neg control				1.924		100.0
Mean % Viability at 3 min:						85.2
Mean % Viability at 60 min:						100.1
Predicted Corrosivity:			Non-corrosive

 

Negative Control:		Tissue Culture Water
Dose:		50µl
Conc:		Neat
TIME (min)	OD 1	OD 2	OD 3	MEAN (OD)	SD	VIABILITY %	ERROR %
3.0	2.026	2.078	2.066	2.057	0.027	100.0	1.3
3.0	2.00	2.073	2.094	2.056	0.049	100.0	2.4
Mean				2.056		100.0
60.0	1.959	1.936	1.961	1.952	0.014	101.4	0.7
60.0	1.859	1.932	1.899	1.897	0.037	98.6	1.9
Mean				1.924		100.0
Predicted Corrosivity:			Non-corrosive

 

Positive Control:		Potassium Hydroxide, 8.0 N
Dose:		50µl
Conc:		Neat
TIME (min)	OD 1	OD 2	OD 3	MEAN (OD)	SD	VIABILITY %	ERROR %
3.0	0.587	0.602	0.602	0.597	0.009	29.0	0.4
3.0	0.705	0.708	0.708	0.707	0.002	34.4	0.1
Neg control				2.056		100.0
60.0	0.038	0.039	0.040	0.039	0.001	2.0	0.1
60.0	0.043	0.043	0.043	0.043	0.000	2.2	0.0
Neg control				1.924		100.0
Mean % Viability at 3 min:						31.7
Mean % Viability at 60 min:						2.1
Predicted Corrosivity:			Corrosive

 

Interpretation of results:

GHS criteria not met

Remarks:

Criteria used for interpretation of results: EU

Conclusions:

H2925 (CAS No. 68130-53-0), Lot# 2013090401 is predicted to be non-corrosive.

Executive summary:

OBJECTIVE: To predict and classify the skin corrosivity potential of a chemical by using a three-dimensional human epidermis model. This study is designed based on MatTek protocol "In Vitro EpiDerm™ Skin Corrosion Test" and follows the OECD Guideline for the Testing of Chemicals No. 431 In Vitro Skin Corrosion: Human Skin Model Test.

METHOD SYNOPSIS: MatTek EpiDerm^TMtissues were treated in duplicate with the test article. Negative Control and Positive Control for 3 minutes and 60 minutes. Following treatment, the viability of thetissues was determined using Methyl thiazole tetrazolium (MTT) uptake and conversion, and theabsorbance of each tissue was measured at 540 nm. The mean viability was then expressed as apercent of control values. The percent viability was used to determine corrosivity potential.

 

SUMMARY/CONCLUSION:

Test and Control Article Identity	Mean Viability (3 min.)	Mean Viability (60 min.)	Predicted Corrosivity
H2925 (CAS No. 68130-53-0), Lot# 2013090401	85.2%	100.1%	Non-Corrosive
Tissue culture water (Negative Control)	100.0%	100.0%	Non-Corrosive
Potassium Hydroxide, 8.0 N (Positive Control)	31.7%	2.1%	Corrosive

 

Reference 6

Endpoint:

skin 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:

See section 13 for the full attached justification for read-across

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Irritation parameter:

erythema score

Basis:

mean

Remarks:

of all three animals

Time point:

other: 24 h, 48 h and 72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility: not applicable

Remarks on result:

other: no cutaneous effects observed

Remarks:

CAS 11138-60-6

Irritation parameter:

edema score

Basis:

mean

Remarks:

of all three animals

Time point:

other: 24 h, 48 h and 72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility: not applicable

Remarks on result:

other: no cutaneous effects observed

Remarks:

CAS 11138-60-6

Irritation parameter:

erythema score

Basis:

mean

Remarks:

out of all 5 animals

Time point:

other: mean over 24, 48, 72 h

Score:

0.24

Max. score:

4

Reversibility:

fully reversible within: 72 h

Remarks on result:

other: CAS 91050-89-4

Irritation parameter:

edema score

Basis:

mean

Remarks:

out of all five animals

Time point:

other: mean over 24, 48, 72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Remarks on result:

other: CAS 91050-89-4

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.66

Max. score:

4

Reversibility:

fully reversible within: 72 h

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

edema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

edema score

Basis:

animal #2

Time point:

24/48 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

edema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 h

Remarks on result:

other: CAS 78-16-0

Interpretation of results:

GHS criteria not met

Conclusions:

The substance, CAS 68130-51-8; EC 268-594-6, is analogous to the substances to be read across to, in terms of basic form, and the degree of substitution of functional groups is not considered to effect the proposed read across for in viivo skin irritation. The substance is not considered to be corrosive or irritating on the basis of read across.

Reference 7

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03 April 2014 to 02 July 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed in accordance with OECD test guidelines in compliance with GLP.

Qualifier:

according to guideline

Guideline:

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

Qualifier:

according to guideline

Guideline:

other: EPA 40 CFR 158.340

GLP compliance:

yes

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: Not specified

Source strain:

not specified

Details on animal used as source of test system:

EpiDerm™ tissues, Lot 19981 Kit D, were received from MatTek Corporation (Ashland, MA) on 08 Apr 2014 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.

Justification for test system used:

In accordance with test guidelines.

Vehicle:

unchanged (no vehicle)

Details on test system:

Test Article Reduction of MIT: 100 µI of the test article were mixed with 1 ml of MTT 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 (TCH2O), 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 MTT Since tissue viability is based on MTT 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 MTT and the assay continued as per the protocol.

Mesh Compatibility: Pre-cut nylon meshes supplied with the tissues were placed on a slide and 30 µI of the test article or TCH2O Negative Control were applied. After 60 minutes exposure, the mesh was checked microscopically, No interaction between the test article or TCH2O and the mesh was observed so the test article was dosed using the mesh as a spreading aid,

Dosing:
30 µI of the test article or control articles were applied to the EpiDerm ™ tissue. The nylon mesh was then placed on top to facilitate even distribution of the test material. A Negative Control (phosphate buffered saline (PBS)) and a Positive Control (5% sodium dodecyl sulfate (SDS) solution, MatTek) were tested concurrently, with a nylon mesh placed on top to facilitate even distribution of the material. Each treatment with test article or control was conducted in triplicate.
The exposure period for the test article and controls was 60 minutes. The dosed tissues were placed in an incubator at 37° ± 1°C, 5% ± 1% CO2 for 35 ± 1 minutes, then returned to the sterile hood for the remainder of the 60-minute exposure period.
After dosing and incubation, the tissues were rinsed with PBS, blotted to remove the test substance and dry the tissue, and transferred to fresh medium. The rinsed EpiDerm™ tissues were returned to the incubator for 24 ± 2 hours. Medium was changed at 24 ± 2 hours. Tissues were returned to the incubator for an additional 18 ± 2 hours.

Tissue Viability (MTT Reduction): At the end of the incubation period, each EpiDerm™ tissue was rinsed with phosphate buffered saline (PBS) and transferred to a 24-well plate containing 300 µI of MTT solution (1 mg/ml MTT in DMEM). The tissues were then returned to the incubator for a three-hour MTT incubation period. Following the MTT incubation period, each EpiDerm TM tissue was rinsed with PBS and then treated with 2.0 ml of extractant solution (isopropanol) per well for at least two hours, with shaking, at room temperature. Two aliquots of the extracted MTT formazan were measured at 540 nm using a plate reader (µQuant Plate Reader, BioTek Instruments, Winooski, VT).

Analysis of Data: The mean absorbance value for each time point was calculated from the optical density (OD) of the duplicate samples and expressed as percent viability for each sample using the following formula:
% viability =100 X (OD sample/OD Negative Control)

Quality Controls: The assay meets the acceptance criterion if the mean OD540 of the Negative Control tissues is ≥ 1.0 and ≤ 2.5, and the mean viability of Positive Control tissues, expressed as percentage of the Negative Control tissues, is ≤ 20%. In addition, the standard deviation (SD) calculated from individual percent tissue viabilities of the three identically-treated replicates is < 18%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

30 µI of the test article were applied to the top of each EpiDerm™ tissue.

Duration of treatment / exposure:

60 minutes

Duration of post-treatment incubation (if applicable):

Not specified

Number of replicates:

Tests were repeated in triplicate.

Irritation / corrosion parameter:

% tissue viability

Value:

96.4

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other:

Remarks:

Basis: mean. Time point: 60 minutes. Max. score: 100.0. Remarks: Viability score are reported as %. The lower the score the more harmful a substance is expected to be. . (migrated information)

Interpretation of results:

GHS criteria not met

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

H2925 (CAS No. 68130-53-0), Lot# 2013090401 is considered to be a non-irritant and therefore it is not classified in accordance with the CLP Regulation.

Executive summary:

OBJECTIVE: To predict dermal irritation potential of test articles in the context of identification and classification of skin irritation hazard according to the European Union (EU) classification, United Nations Globally Harmonized System of Classification and Labelling of Chemicals (GHS) classification system (Category 2 and non-irritants), and OECD Guideline for the Testing of Chemicals No. 439 - In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method. This study is designed based on MatTek protocol in vitro EpiDerm™ Skin Irritation Test.

METHOD SYNOPSIS: MatTek EpiDerm™ tissue samples were treated in triplicate with the test article, Negative Control and Positive Control for 60 minutes. Following treatment and subsequent incubation time, the viability of the tissues was determined using Methylthiazole tetrazolium (MTT) uptake and reduction. The absorbance of each sample was measured at 540 nm. The viability was then expressed as a percent of control values. If the mean tissue viability was ≤ 50%, the test material was classified as an irritant; if the mean tissue Viability was >50%, the test material was classified as a non-irritant.

SUMMARY/CONCLUSION:

Test and Control Article Identity	Mean Tissue Viability	Irritancy Classification
H2925 (CAS No. 68130-53-0), Lot# 2013090401	96.4%	Non-Irritant
Phosphate Buffered Saline (Negative Control)	100.0%	Non-Irritant
5% Sodium Dodecyl Sulfate (Positive Control)	2.4%	Irritant

 

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not 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:

31. Aug. 2017

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)

Version / remarks:

OECD Guideline for the Testing of Chemicals, Method No. 437, edition adopted 26. Jul. 2013: “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)

Version / remarks:

Commission Regulation (EU) 2017/735 amending Regulation (EC) No. 440/2008, EU Method B.47 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, adopted 14. Feb. 2017

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

No further details specified in the study report.

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

Specification
Species Bos primigenius Taurus (fresh bovine corneas)

Origin
Fresh bovine eyes were obtained from the slaughterhouse Müller Fleisch GmbH, Enzstr. 2-4, 75217 Birkenfeld, Germany, on the day of the test. The cattle were between 12 and 60 months old. The eyes were transported to the test facility in Hanks’ Balanced Salt Solution with 1% Penicillin-Streptomycin solution (Penicillin 100 U/mL, Streptomycin 100 μg/mL) in a suitable cooled container.

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

750 μL

Duration of treatment / exposure:

10 minutes

Duration of post- treatment incubation (in vitro):

2 hours

Number of animals or in vitro replicates:

For each treatment group (negative control solution, test item and positive control), three replicates were used.

Details on study design:

Preparations
After having carefully cleaned and sterilised the cornea holders, they were kept in the incubation chamber at 32 ± 1 °C.
On the day of the assay, the MEM without phenol red was supplemented with sodium bicarbonate, L-glutamine and 1% fetal calf serum (= complete MEM) and stored in a water bath at 32 ± 1 °C.
The same was performed with the MEM with phenol red, but without addition of sodium bicarbonate.
After the arrival of the corneas, they were examined and only corneas which were free from damages were used. The corneas were excised with a scalpel and cut from the globe with a 2-3 mm ring of sclera around the outside. Each cornea was transferred to a cornea holder in which pre-warmed cMEM (32 ± 1 °C) without phenol red was filled. The holders were then incubated for 1 hour in the incubation chamber at 32 ± 1 °C.

Experimental Parameters
Date of treatment: 31. Aug. 2017
Incubation time: 10 min.
Negative control: HBSS
Positive control: Dimethylformamide (undiluted)

Method Description
After the initial incubation, the medium was changed and the baseline opacity for each cornea was recorded. None of the corneas showed tissue damage; therefore, all corneas were used. For each treatment group (negative control solution, test item and positive control), three replicates were used. After removal of the pre-incubation medium (cMEM without phenol red), 750 μL negative control solution, test item and positive control were applied to each replicate.
According to the characteristics of the test item, the following treatment procedure was performed:

Closed Chamber Method
The respective substance (negative control solution, test item or positive control) was applied by pipetting 750 μL of the appropriate liquid through the refill hole in the holder on the cornea. The test item was given on the epithelium in such a manner that as much as possible of the cornea was covered with the test item.
Exposure time of the test item on the corneas was 10 minutes at 32 ± 1 °C. After thorough rinsing with cMEM with phenol red and final rinsing with cMEM without phenol red, the anterior chamber was filled with cMEM without phenol red, and the corneas were stored for additional 2 hours at 32 ± 1 °C (post-incubation).
After post-incubation time, the cMEM without phenol red was renewed in both chambers.
Then, the final opacity value of each cornea was recorded. The cMEM without phenol red was removed from the front chamber, and 1 mL sodium fluorescein solution (concentration: 4 mg/mL) was added to the front chamber.
The chambers were then closed again and incubated for 90 minutes at 32 ± 1 °C. After incubation, the content of the posterior chamber was thoroughly mixed. Then, the permeability of the cornea was measured as optical density of the liquid with a microtiter plate photometer at 492 nm.

Irritation parameter:

in vitro irritation score

Run / experiment:

Test Item

Value:

0.41

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

The in vitro study was performed to assess corneal damage potential of Hatcol 1570 by quantitative measurements of changes in opacity and permeability in a bovine cornea.
The test item Hatcol 1570 was brought onto the cornea of a bovine eye which previously had been incubated with cMEM without phenol red at 32 ± 1 °C for 1 hour and whose opacity had been determined. The test item was incubated on the cornea for 10 minutes at 32 ± 1 °C. After removal of the test item and 2 hours post-incubation, opacity and permeability values were measured.
The test item was tested pure.
Under the conditions of the test, the test item Hatcol 1570 showed no effects on the cornea of the bovine eye. The calculated IVIS (in vitro irritancy score) is 0.41.
According to OECD Guideline no. 437 (Jul. 2013), a substance with an IVIS ≤ 3 requires no classification for eye irritation or serious eye damage.
The negative control (HBSS) and the positive control (undiluted dimethylformamide) have met the validity criteria.
No observations were made which might cause doubts concerning the validity of the study outcome. The test is considered valid.

Opacity and Permeability Values

The illuminance (unit: LUX) values which were measured before and after exposure are given in the following table:

Illuminance Values

Parameter	Negative Control			Test Item			Positive Control
	1. Rep.	2. Rep.	3. Rep.	1. Rep.	2. Rep.	3. Rep.	1. Rep.	2. Rep.	3. Rep.
(I) Measured values before exposure	997	989	1043	1003	1017	976	1009	990	983
(I) Measured values after exposure	1006	971	1020	981	1001	961	422	431	403

Rep. = Replicate

 

The values in the following tables present the calculated opacity values, according to evaluation:

Opacity Values Negative Control

Parameter	Negative Control
	1. Rep.	2. Rep.	3. Rep.
Opacity before exposure	4.30	4.65	2.37
Opacity after exposure	3.91	5.47	3.31
Opacity Difference	-0.39	0.82	0.94
Mean Opacity Difference	0.46

Rep. = Replicate

 

Opacity Values Test Item and Positive Control

Parameter	Test item			Positive Control
	1. Rep.	2. Rep.	3. Rep.	1. Rep.	2. Rep.	3. Rep.
Opacity before exposure	4.04	3.44	5.24	3.78	4.61	4.92
Opacity after exposure	5.01	4.12	5.93	63.86	61.70	68.73
Opacity Difference	0.97	0.69	0.70	60.08	57.10	63.81
Opacity Difference corrected	0.52	0.23	0.24	59.63	56.64	63.36
Mean Opacity Difference corrected	0.33			59.88

Rep. = Replicate

 

For the permeability measurement, three replicates for each treatment group were measured three time. cMEM without phenol red was measured as blank value as well. The optical density values at 492 nm are given in the following tables:

Optical density at 420 nm of Blank

Parameter	cMEM without phenol red
1. Measurement	0.038
2. Measurement	0.041
3. Measurement	0.041
Mean	0.040

 

Optical density at 420 nm of Negative Control, Test Item and Positive Control

Parameter	Negative Control			Test item			Positive Control
	1. Rep.	2. Rep.	3. Rep.	1. Rep.	2. Rep.	3. Rep.	1. Rep.	2. Rep.	3. Rep.
1. Measurement	0.036	0..039	0.041	0.039	0..048	0.044	0.505	0.605	0.450
2. Measurement	0.034	0.041	0.038	0.044	0.046	0.044	0.509	0.614	0.453
3. Measurement	0.040	0.037	0.040	0.039	0.050	0.043	0.515	0.605	0.450
1. Measurement – blank	-0.0040	-0.0010	0.0010	-0.0010	0.0080	0.0040	0.4650	0.5650	0.4100
2. Measurement – blank	-0.0060	0.0010	-0.0020	0.0040	0.0060	0.0040	0.4690	0.5740	0.4130
3. Measurement – blank	0.0000	-0.0030	0.0000	-0.0010	0.0100	0.0030	0.4750	0.5650	0.4100
Mean of each replicate	-0.0033	-0.0010	-0.0003	0.0007	0.0080	0.0037	0.4697	0.5680	0.4110
Mean of the 3 replicate	-0.0016			--			--
Corrected	--	--	--	0.0022	0.0096	0.0052	0.4712	0.5696	0.4126
Corrected mean of the 3 replicates	--			0.0057			0.4844

Rep. = Replicate

 

IVIS Values

The calculated IVIS for each replicate and the corresponding means are presented in the following table:

IVIS

Test Group	IVIS	Mean IVIS	Relative Standard Deviation IVIS
Negative Control HBSS	-0.44	0.43	175.58%
	0.80
	0.94
Test Item Hatcol 1570	0.55	0.41	23.39%
	0.37
	0.32
Positive Control DMF undiluted	66.70	67.14	3.30%
	65.19
	69.54

Note: the high relative standard deviations of the IVIS of test item and negative control are due to mathematical reasons, as the respective means are very small.

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the study, the test item Hatcol 1570 showed no effects on the cornea of the bovine eye. The calculated IVIS (in vitro irritancy score) is 0.41.
According to OECD Guideline no. 437 (Jul. 2013), a substance with an IVIS ≤ 3 requires no classification for eye irritation or serious eye damage.

Executive summary:

Evaluation of Hatcol 1570 in the Bovine Corneal Opacity and Permeability (BCOP) Test Method following OECD Guideline 437 and EU Method B.47

 

Findings and Results:

One valid experiment was performed.

Bovine corneas were used. They were collected from slaughtered cattle which were between 12 and 60 months old.

The test item Hatcol 1570 was brought onto the cornea of a bovine eye which had been incubated with cMEM without phenol red at 32 ± 1 °C for 1 hour and whose opacity had been measured.

The test item was incubated on the cornea for 10 minutes at 32 ± 1 °C. After removal of the test item and 2 hours post-incubation, opacity and permeability values were measured.

 

HBSS was used as negative control. The negative control showed no irritating effect on the cornea and the calculated IVIS (in vitro irritancy score) is 0.43.

Dimethylformamide (DMF) undiluted was used as positive control. The positive control induced serious eye damage on the cornea and was within two standard deviations of the current historical mean. The calculated IVIS (in vitro irritancy score) is 67.14.

 

Under the conditions of the study, the test item Hatcol 1570 showed no effects on the cornea of the bovine eye. The calculated IVIS (in vitro irritancy score) is 0.41.

 

According to OECD Guideline no. 437 (Jul. 2013), a substance with an IVIS ≤ 3 requires no classification for eye irritation or serious eye damage.

Reference 2

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions. Limited documentation but relevant data given.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

Adopted 24 April 2002

Deviations:

yes

Remarks:

limited documentation, limited information on test substance given

GLP compliance:

not specified

Species:

rabbit

Strain:

New Zealand White

Vehicle:

unchanged (no vehicle)

Controls:

other: the untreated eye served as control

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.1 mL
- Concentration: 100 %

Duration of treatment / exposure:

Group 1: 10 s
Group 2: 72 h (continuous treatment)

Observation period (in vivo):

72 h
Reading time points: 1, 24, 48 and 72 h

Number of animals or in vitro replicates:

Group 1: 4 males
Group 2: 4 males

Details on study design:

REMOVAL OF TEST SUBSTANCE
Group 1
- Washing: yes
- Time after start of exposure: 10 s
Group 2
- Washing: no

SCORING SYSTEM: Draize scoring system

TOOL USED TO ASSESS SCORE: fluorescein

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

out of all 4 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Irritation parameter:

iris score

Basis:

mean

Remarks:

out of all 4 animals

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: reversibility not applicable

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

out of all 4 animals

Time point:

24/48/72 h

Score:

0

Max. score:

3

Reversibility:

other: reversibility not applicable

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

out of all 4 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Irritant / corrosive response data:

No effects on cornea and iris were observed. Slight redness of the conjunctivae was observed both, after 10 s and after continuous treatment. The effects were obvious after 1 and 6 h. After 10 s treatment, the effects were reversible within 48 h. After continuous treatment, redness reversed within 24 h. In addition, exsudation was observed in both treatment groups, being reversible within 24 h (continuous treatment) and 48 h (10 s treatment), respectively. Chemosis was not observed.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Reference 3

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

Adopted: 24 April 2002

Deviations:

yes

Remarks:

limited information on test substance

GLP compliance:

not specified

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Weight at study initiation: 3408 - 3935 g

Vehicle:

unchanged (no vehicle)

Controls:

other: the untreated eye served as control

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.1 mL
- Concentration: 100 %

Duration of treatment / exposure:

5 days (single application without washing)

Observation period (in vivo):

5 days
Reading time points: 2, 24, 48 and 72 h

Number of animals or in vitro replicates:

3 males

Details on study design:

REMOVAL OF TEST SUBSTANCE: no

SCORING SYSTEM: Draize scoring system

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

out of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Irritation parameter:

iris score

Basis:

mean

Remarks:

out of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: reversibility not applicable

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

out of all 3 animals

Time point:

24/48/72 h

Score:

0.3

Max. score:

3

Reversibility:

fully reversible within: 48 h

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

out of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible within: reversibility not applicable

Irritant / corrosive response data:

No effects on the cornea and the iris were observed. Conjunctival effects included slight to moderate redness and slight to severe discharge. No chemosis was seen. Additional signs of irritation included erythema and dried secretion on the upper and lower eyelids and convoluted eyelids. All effects were completely reversible within 48 h after application.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Reference 4

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restrictions.

Qualifier:

equivalent or similar to guideline

Guideline:

other: 16 CFR 1500.42

Deviations:

yes

Remarks:

limited documentation

GLP compliance:

no

Species:

rabbit

Strain:

other: albino rabbit, not further specified

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Acclimation period was carried out, not further specified.

Vehicle:

unchanged (no vehicle)

Controls:

other: the untreated eye served as control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL

Duration of treatment / exposure:

single application without washing

Observation period (in vivo):

72 h
Reading time points: 24, 48 and 72 h

Number of animals or in vitro replicates:

6

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: no washing was done

SCORING SYSTEM: Draize scoring system

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

out of all 6 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility: not applicable

Irritation parameter:

iris score

Basis:

mean

Remarks:

out of all 6 animals

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: reversibility: not applicable

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

out of all 6 animals

Time point:

24/48/72 h

Score:

0.4

Max. score:

3

Reversibility:

fully reversible within: 72 h

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

out of all 6 animals

Time point:

24/48/72 h

Score:

0.1

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritant / corrosive response data:

No effects on the cornea or iris were observed in test animals. Slight redness of the conjunctivae was observed after 24 h in 3 animals and after 48 h in 4 animals which were fully reversible in all animals within 72 h.
Slight chemosis of the conjunctiva was detected in 1 animal after 24 h which was fully reversible within 48 h.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Reference 5

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:

See section 13 for the full attached justification for read-across

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

GLP compliance:

yes

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

3

Reversibility:

other: reversibility: not applicable

Remarks on result:

other: only slight exsudation at 1h (2 cases) and 24 h (1 case) was seen

Remarks:

CAS 11138-60-6

Irritation parameter:

iris score

Basis:

mean

Remarks:

of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: reversibility: not applicable

Remarks on result:

other: none of the animals showed effects in the iris

Remarks:

CAS 11138-60-6

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Remarks on result:

other: none of the aniamals showed corneal opacity

Remarks:

CAS 11138-60-6

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Remarks on result:

other: none of the animals showed chemosis

Remarks:

CAS 11138-60-6

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

out of all 6 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

iris score

Basis:

mean

Remarks:

out of all 6 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0

Max. score:

2

Reversibility:

other:

Remarks on result:

other: not applicable

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

out of all 6 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0.4

Max. score:

3

Reversibility:

fully reversible within: 72 h

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

out of all 6 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0.1

Max. score:

4

Reversibility:

fully reversible within: 48 h

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

out of all 4 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Remarks on result:

other: CAS 91050-89-4

Irritation parameter:

iris score

Basis:

mean

Remarks:

out of all 4 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0

Max. score:

2

Reversibility:

other: reversibility not applicable

Remarks on result:

other: CAS 91050-89-4

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

out of all 4 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0

Max. score:

3

Reversibility:

other: not applicable

Remarks on result:

other: CAS 91050-89-4

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

out of all 4 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Remarks on result:

other: CAS 91050-89-4

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

out of all 3 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

iris score

Basis:

mean

Remarks:

out of all 3 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0

Max. score:

2

Reversibility:

other: reversibility not applicable

Remarks on result:

other: CAS 78-16-0

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

out of all 3 animals

Time point:

other: mean over 24, 48 and 72 h

Score:

0.3

Max. score:

3

Reversibility:

fully reversible within: 48 h

Remarks on result:

other: CAS 78-16-0

Interpretation of results:

not irritating

Remarks:

Based on read-across

Conclusions:

The substance, CAS 68130-51-8; EC 268-594-6, is analogous to the substances to be read across to, in terms of basic form, and the degree of substitution of functional groups is not considered to effect the proposed read across for the endpoint of in vivo eye irritation. The substance is not considered to be irritating or corrosive to eyes on the basis of read across.

Reference 6

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20 Nov - 23 Nov 1990

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP guideline study; only few details on study substance). For read-across, maximum reliability score is 2.

Justification for type of information:

GLP guideline study; only few details on study substance). For read-across, maximum reliability score is 2.

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

adopted in 1987

Deviations:

yes

Remarks:

lack of details on test substance

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Elevage Cunicole du Val de Selle, France
- Weight at study initiation: 2.6 kg
- Housing: individually
- Diet: Certified pellet (Rabbits sustenance ref. 112 C; ad libitum
- Water: tap water, ad libitum
- Acclimation period: 5 days
-other: the animals received a preventive coccidiosis treatment during acclimation (Mucoxid, 137.5 mg/kg bw/day, via drinking water)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17-23
- Humidity (%): 30-70
- Photoperiod (hrs dark / hrs light): 12 / 12

Vehicle:

unchanged (no vehicle)

Controls:

other: the untreated eye served as control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL

Duration of treatment / exposure:

single dose, no rinsing

Observation period (in vivo):

72 h
Reading time points: 1, 24, 48 and 72 h

Number of animals or in vitro replicates:

3 males

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): No

SCORING SYSTEM: according to 84/499/EEC appendix V B5, i.e., according to the Draize scoring system

TOOL USED TO ASSESS SCORE: fluorescein

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

3

Reversibility:

other: reversibility: not applicable

Remarks on result:

other: only slight exsudation at 1h (2 cases) and 24 h (1 case) was seen

Irritation parameter:

iris score

Basis:

mean

Remarks:

of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: reversibility: not applicable

Remarks on result:

other: none of the animals showed effects in the iris

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Remarks on result:

other: none of the aniamals showed corneal opacity

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

of all 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: reversibility not applicable

Remarks on result:

other: none of the animals showed chemosis

Irritant / corrosive response data:

In 2 animals slight exsudation (grade 1) was obsevered at 1 h. At 24 h slight exsudation (grade 1) was observed in 1 animals. These effects were completely reversible at 48 h.

Animal	Hours after application
	1					24					48					72
	A	B	C	D	E	A	B	C	D	E	A	B	C	D	E	A	B	C	D	E
1	0	0	0	0	1	0	0	0	0	1	0	0	0	0	0	0	0	0	0	0
2	0	0	0	0	1	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
3	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0

A = Cornea Score

B = Iris Score

C = Erythema Score

D = Chemosis Score

E = Exsudation Score

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin Corrosion/Irritation

In vivo skin corrosion/irritation:

Three in vivo skin irritation studies are also available for read-across substances:

The first study was conducted with CAS 11138-60-6 in accordance with OECD Guideline 404. No cutaneous reactions occurred in the 3 New Zealand White rabbits tested. Therefore, it was concluded that the test substance is not irritating to the skin.

The second study was conducted with CAS 91050-89-4 in accordance with OECD Guideline 404. After 24 h, 3 animals (New Zealand White rabbits) showed erythema, which persisted in 1 animal for 48 h. 72 h after exposure, all animals were free of erythema. Therefore, it was concluded that the test substance is not irritating to the skin.

The third study was conducted with CAS 78-16-0 in accordance with EU Method B.4. Very slight erythema and edema were observed in all animals (3 New Zealand White rabbits) 24 h after treatment which were fully reversible latest within 72 h. Therefore, it was concluded that the test substance is not irritating to the skin.

In vitro skin irritation:

GLP studies conducted according OECD Guideline 431 and 439 using reconstructed human epidermis concluded that the CAS No. 68130-53-0 is not irritating or corrosive to skin.

Eye Damage/Irritation

In vitro Eye Irritation: Bovine Corneal Opacity and Permeability (BCOP) Test

Bovine corneas were used. They were collected from slaughtered cattle which were between 12 and 60 months old.

The test item Hatcol 1570 was brought onto the cornea of a bovine eye which had been incubated with cMEM without phenol red at 32 ± 1 °C for 1 hour and whose opacity had been measured.

The test item was incubated on the cornea for 10 minutes at 32 ± 1 °C. After removal of the test item and 2 hours post-incubation, opacity and permeability values were measured.

HBSS was used as negative control. The negative control showed no irritating effect on the cornea and the calculated IVIS (in vitro irritancy score) is 0.43.

Dimethylformamide (DMF) undiluted was used as positive control. The positive control induced serious eye damage on the cornea and was within two standard deviations of the current historical mean. The calculated IVIS (in vitro irritancy score) is 67.14.

 

Under the conditions of the study, the test item Hatcol 1570 showed no effects on the cornea of the bovine eye. The calculated IVIS (in vitro irritancy score) is 0.41.

According to OECD Guideline no. 437 (Jul. 2013), a substance with an IVIS ≤ 3 requires no classification for eye irritation or serious eye damage.

In vivo eye irritation

Four in vivo eye irritation studies are available for read-across substances:

The first study was conducted with CAS 11138-60-6 in accordance with OECD Guideline 405. In 2 animals (New Zealand White rabbits) slight exudation (grade 1) was observed at 1 h. At 24 h slight exudation (grade 1) was observed in 1 animals. These effects were completely reversible at 48 h. Therefore, it was concluded that the test substance is not irritating to the eyes.

The second study was conducted with CAS 78-16-0 in accordance with guideline 16 CFR 1500.42. No effects on the cornea or iris were observed in test animals. Slight redness of the conjunctivae was observed after 24 h in 3 animals (albino rabbits) and after 48 h in 4 animals which were fully reversible in all animals within 72 h. Slight chemosis of the conjunctiva was detected in 1 animal after 24 h which was fully reversible within 48 h.Therefore, it was concluded that the test substance is not irritating to the eyes.

The third study was conducted with CAS 91050-89-4 in accordance with OECD Guideline 405. No effects on cornea and iris were observed. Slight redness of the conjunctivae was observed both, after 10 s and after continuous treatment. The effects were obvious after 1 and 6 h. After 10 s treatment, the effects were reversible within 48 h. After continuous treatment, redness reversed within 24 h. In addition, exudation was observed in both treatment groups, being reversible within 24 h (continuous treatment) and 48 h (10 s treatment), respectively. Chemosis was not observed. Therefore, it was concluded that the test substance is not irritating to the eyes.

The fourth study was conducted with CAS 78-16-0 in accordance with OECD Guideline 405 in New Zealand White rabbits. No effects on the cornea and the iris were observed. Conjunctival effects included slight to moderate redness and slight to severe discharge. No chemosis was seen. Additional signs of irritation included erythema and dried secretion on the upper and lower eyelids and convoluted eyelids. All effects were completely reversible within 48 h after application. Therefore, it was concluded that the test substance is not irritating to the eyes.

Justification for classification or non-classification

In accordance with Regulation (EC) 1272/2008, the available information does not allow for classification based on the lack of effects.

A report justifying the read-across approach is included in IUCLID Chapter 13.