Registration Dossier
Registration Dossier
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EC number: 604-580-1 | CAS number: 147256-33-5
Endpoint summary
Administrative data
Description of key information
Skin irritation (OECD 404): not irritating
Eye irritation (OECD 405) not irritating
Key value for chemical safety assessment
Skin irritation / corrosion
Link to relevant study records
- Endpoint:
- skin irritation: in vivo
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 06 - 16 Dec 2011
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: GLP guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- The department of health of the government of the United Kingdom
- Species:
- rabbit
- Strain:
- New Zealand White
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Strain (as specified by author): Hsdlf:NZW
- Source: Harlan Laboratories UK Ltd., Leicestershire, UK
- Age at study initiation: 12-20 weeks
- Weight at study initiation: 2.40-2.84 kg
- Housing: individually in suspended cages with environmental enrichment items
- Diet: ad libitum, 2930 Teklad Global Rabbit diet (Harlan Laboratories Ltd., Oxon, UK)
- Water : ad libitum, mains drinking water
- Acclimatisation: at least 5 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17-23
- Humidity (%): 30-70
- Air changes (per hr): at least 15
- Photoperiod (hrs dark / hrs light): 12/12 - Type of coverage:
- semiocclusive
- Preparation of test site:
- other: clipped
- Vehicle:
- unchanged (no vehicle)
- Controls:
- not required
- 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: immediately after patch removal, 1, 24, 48 and 72 h - Number of animals:
- 3 males
- Details on study design:
- TEST SITE
- Area of exposure: 2.5 cm x 2.5 cm of the back of the rabbit
- Type of wrap if used: cotton gauze patch secured in position with a strip of surgical adhesive tape, the trunk of the rabbit was wrapped in an elasticated corset during exposure
REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes, any residual test item was removed by gentle swabbing with cotton wool saked in distilled water
- Time after start of exposure: 4h
SCORING SYSTEM: according to Draize scoring system - Irritation parameter:
- erythema score
- Basis:
- animal: #1
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 4
- Reversibility:
- other: reversibility not applicable
- Irritation parameter:
- erythema score
- Basis:
- animal: #2
- Time point:
- 24/48/72 h
- Score:
- 0.67
- Max. score:
- 4
- Reversibility:
- fully reversible within: 72 h
- Irritation parameter:
- erythema score
- Basis:
- animal: #3
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 4
- Reversibility:
- other: reversibility not applicable
- Irritation parameter:
- edema score
- Basis:
- mean
- Remarks:
- out of all three animals
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 4
- Reversibility:
- other: reversibility not applicable
- Irritant / corrosive response data:
- Very slight erythema (grade 1) was noted at one treated skin site at the 24 and 48 h reading time points. Effects were fully reversible within 72 h.
- Other effects:
- All animals showed expected gain in bodyweight during the study.
- Interpretation of results:
- not irritating
- Remarks:
- Migrated information Criteria used for interpretation of results: EU
- Conclusions:
- CLP: not classified
DSD: not classified - Endpoint:
- skin irritation: in vivo
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 19 June - 27 June 1989
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Guideline study conducted according to GLP with acceptable restrictions: Limited details on test substance given.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
- Version / remarks:
- adopted in 1981
- Deviations:
- yes
- Remarks:
- Limited details on test substance
- GLP compliance:
- yes
- Species:
- rabbit
- Strain:
- other: Kleinrusse, Chbb:HM
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Fa. Thomae, Biberach, Germany
- Age at study initiation: 5-6 months
- Weight at study initiation: 2590 g (mean)
- Diet: Altromin 2023, ad libitum
- Water: tap water, ad libitum
- Acclimatisation: at least 1 week
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-25
- Humidity (%): 45-70
- Photoperiod (hrs dark / hrs light): 12/12 - Type of coverage:
- semiocclusive
- Preparation of test site:
- shaved
- Vehicle:
- unchanged (no vehicle)
- Controls:
- other: not required, untreated site 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:
- 7 d
reading time points: 1, 24, 48 and 72 h - Number of animals:
- 3
- Details on study design:
- TEST SITE
- Area of exposure: 6 cm²
SCORING SYSTEM: according to the Draize scoring system - Irritation parameter:
- erythema score
- Basis:
- animal #1
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 4
- Reversibility:
- other: reversibility: not applicable
- Irritation parameter:
- erythema score
- Basis:
- animal #2
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 4
- Reversibility:
- other: reversibility: not applicable
- Irritation parameter:
- erythema score
- Basis:
- animal #3
- Time point:
- 24/48/72 h
- Score:
- 1
- Max. score:
- 4
- Reversibility:
- fully reversible within: 7 d
- Irritation parameter:
- edema score
- Basis:
- mean
- Remarks:
- out of all three animals
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 4
- Reversibility:
- other: reversibility: not applicable
- Irritant / corrosive response data:
- One animal showed erythema graded 1 at each of the reading time points 24, 48 and 72 h, but no edema; full reversibility was achieved on day 7 of observation. The two remaining animals showed no skin reaction at all.
- Interpretation of results:
- not irritating
- Remarks:
- Migrated information Criteria used for interpretation of results: EU
- Conclusions:
- CLP: not classified
DSD: not classified - Endpoint:
- skin corrosion: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Guideline study; limited documentation
- Qualifier:
- according to guideline
- Guideline:
- other: OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)
- Deviations:
- yes
- Remarks:
- (limited documentation)
- GLP compliance:
- not specified
- Remarks:
- the limited information in the study report did not include a confirmation of the GLP status
- Test system:
- human skin model
- Source species:
- human
- Cell type:
- non-transformed keratinocytes
- Cell source:
- other: EpiSkin model
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- TEST SYSTEM
The test consists of a topical exposure of the test substance to the surface of Reconstituted human epidermis model (EpiDermTM) or 3 minutes and 1 hour followed by a cell viability test. Cell viabiltiy is measured by dehydrgenase conversion of the yellow, water-soluble MTT (3-[4,5-dimethylthazol-2-yl]-2,5-diphenyltetrazolium bromide) present in cell mitocondria, in to a blue formazan salt that is measured quantitatively after isopporpanaol exraction from the tissues. The optical density of the extracts of test substance treated tissues is compared to negative ocntrol values from tissue treated with de-ionized water or PBS and is expressed as relatvie tissue viability. - Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- Amount/concentration applied:
- No data
- Duration of treatment / exposure:
- 3 min and 1 hour
- Number of replicates:
- 2 tissues per treatment time
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 1
- Value:
- 92
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Other effects / acceptance of results:
- Mean viability: 92% after 3 minutes (positive control: 13%); 100% after 1 hour (positive control: 11%)
- Interpretation of results:
- other: not corrosive
- Conclusions:
- There is regulatory acceptance in the EU that a substance can be considered corrosive based on a positive result in one of the available in vitro test methods, e.g. the human skin model (HSM) test (OECD 431). Negative in vitro corrosivity responses are not conclusive with respect to non-classification or classification as irritant (Category 2/R37) and shall therefore be subject to further evaluation.
- Endpoint:
- skin irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Guideline study; limited documentation
- Qualifier:
- according to guideline
- Guideline:
- other: OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Deviations:
- yes
- Remarks:
- (limited documentation)
- GLP compliance:
- not specified
- Remarks:
- the limited information in the study report did not include a confirmation of the GLP status
- Test system:
- human skin model
- Source species:
- human
- Cell type:
- non-transformed keratinocytes
- Cell source:
- other: EpiSkin model
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- The test consists of a topical exposure of the test substance to the surface of Reconstituted human epidermis model (EpiDermTM) for 1 hour with 42 hours postincubation followed by a cell viability test. Cell viability is measured by dehydrogenase conversion of the yellow, water-soluble MTT (3-[4,5-dimethylthazol-2-yl]-2,5-diphenyltetrazolium bromide) present in cell mitochondria, in to a blue formazan salt that is measured quantitatively after isopropanol extraction from the tissues. The optical density of the extracts of test substance treated tissues is compared to negative control values from tissue treated with PBS and is expressed as relative tissue viability.
- Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- Amount/concentration applied:
- No data
- Duration of treatment / exposure:
- 1 hour
- Number of replicates:
- 3 tissues per condition
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 1
- Value:
- 112
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Other effects / acceptance of results:
- Mean cell viability: 112% (positive control: 6%)
- Interpretation of results:
- not classified
- Remarks:
- Migrated information Criteria used for interpretation of results: EU
- Conclusions:
- CLP: not classified
DSD: not classified
Referenceopen allclose all
Table 1: Erythema score
Animal Number |
1 h |
1 day |
2 days |
3 days |
71416 |
0 |
0 |
0 |
0 |
71453 |
0 |
1 |
1 |
0 |
71454 |
0 |
0 |
0 |
0 |
Table 2: Edema score
Animal Number |
1 h |
1 day |
2 days |
3 days |
71416 |
0 |
0 |
0 |
0 |
71453 |
0 |
0 |
0 |
0 |
71454 |
0 |
0 |
0 |
0 |
Table 1: Erythema score
Animal Number |
1 h |
24 h |
48 h |
72 h |
7 days |
1168 |
0 |
0 |
0 |
0 |
0 |
1170 |
0 |
0 |
0 |
0 |
0 |
1175 |
0 |
1 |
1 |
1 |
0 |
Table 2: Edema Score
Animal Number |
1 h |
24 h |
48 h |
72 h |
1168 |
0 |
0 |
0 |
0 |
1170 |
0 |
0 |
0 |
0 |
1175 |
0 |
0 |
0 |
0 |
Table 1: Results of the Skin Corrosion Test
Test substance |
|
Exposure 3 min |
Exposure 1 hour |
||||
|
Tissue 1 |
Tissue 2 |
Mean |
Tissue 1 |
Tissue 2 |
mean |
|
NC |
Mean OD570 |
1.851 |
1.889 |
1.870 |
1.641 |
1.777 |
1.709 |
Viability [% of NC] |
99.0 |
101.0 |
100 |
96.0 |
104.0 |
100 |
|
Test item |
Mean OD570 |
1.559 |
1.872 |
1.715 |
1.700 |
1.719 |
1.710 |
Viability [% of NC] |
83.4 |
100.1 |
92 |
99.5 |
100.6 |
100 |
|
PC |
Mean OD570 |
0.235 |
0.254 |
0.244 |
0.229 |
0.155 |
0.192 |
Viability [% of NC] |
12.6 |
13.6 |
13 |
13.4 |
9.0 |
11 |
|
NC = negative control
PC = positive control
OD570 = Optical Density [wavelength 570 nm]
Table 1: Results of the Skin Irritation Test
Test substance |
|
Tissue 1 |
Tissue 2 |
Tissue 3 |
Mean |
SD |
NC |
Mean OD570 |
1.845 |
1.541 |
1.747 |
1.711 |
|
Viability [% of NC] |
107.8 |
90.1 |
102.1 |
100 |
9.06 |
|
Test item |
Mean OD570 |
1.959 |
1.742 |
2.040 |
1.914 |
|
Viability [% of NC] |
114.5 |
101.8 |
119.2 |
112 |
9.02 |
|
PC |
Mean OD570 |
0.092 |
0.098 |
0.094 |
0.095 |
|
Viability [% of NC] |
5.4 |
5.7 |
5.5 |
6 |
0.19 |
NC = negative control
PC = positive control
OD570 = Optical Density [wavelength 570 nm]
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (not irritating)
Eye irritation
Link to relevant study records
- Endpoint:
- eye irritation: in vivo
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 22 Dec 2011- 07 Jan 2012
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: GLP guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 405 (Acute Eye Irritation / Corrosion)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- The department of health of the government of the United Kingdom
- Species:
- rabbit
- Strain:
- New Zealand White
- Details on test animals or tissues and environmental conditions:
- TEST ANIMALS
- Strain (as specified by author): Hsdlf:NZW
- Source: Harlan Laboratories UK Ltd., Leicestershire, UK
- Age at study initiation: 12-20 weeks
- Weight at study initiation: 2.40-2.84 kg
- Housing: individually in suspended cages with environmental enrichment items
- Diet: ad libitum, 2930 Teklad Global Rabbit diet (Harlan Laboratories Ltd., Oxon, UK)
- Water: ad libitum, mains drinking water
- Acclimatisation: at least 5 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17-23
- Humidity (%): 30-70
- Air changes (per hr): at least 15
- Photoperiod (hrs dark / hrs light): 12/12 - Vehicle:
- unchanged (no vehicle)
- Controls:
- other: left 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: 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): Yes, eye was rinsed with water
- Time after start of exposure: 24 h
SCORING SYSTEM: Draize scoring system
TOOL USED TO ASSESS SCORE: standard opthalmoscope - Irritation parameter:
- cornea opacity score
- Basis:
- mean
- Remarks:
- out of all three 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 three animals
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 2
- Reversibility:
- other: reversibility: not applicable
- Irritation parameter:
- conjunctivae score
- Basis:
- animal #1
- Time point:
- 24/48/72 h
- Score:
- 0.33
- Max. score:
- 3
- Reversibility:
- fully reversible within: 48 h
- Irritation parameter:
- conjunctivae score
- Basis:
- animal #2
- Time point:
- 24/48/72 h
- Score:
- 0.33
- Max. score:
- 3
- Reversibility:
- fully reversible within: 48 h
- Irritation parameter:
- conjunctivae score
- Basis:
- animal #3
- Time point:
- 24/48/72 h
- Score:
- 0.33
- Max. score:
- 3
- Reversibility:
- fully reversible within: 48 h
- Irritation parameter:
- chemosis score
- Basis:
- mean
- Remarks:
- out of all three animals
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 4
- Reversibility:
- other: reversibility: not applicable
- Irritant / corrosive response data:
- No corneal or iridial effects were noted during the study. Minimal conjunctival irritation (grade 1) was noted in all treated eyes 1 and 24 h after test substance instillation. Effects were fully reversible within 48 h.
- Other effects:
- All animals showed expected gain in bodyweight during the study.
- Interpretation of results:
- not irritating
- Remarks:
- Migrated information Criteria used for interpretation of results: EU
- Conclusions:
- CLP: not classified
DSD: not classified - Endpoint:
- eye irritation: in vivo
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 03 July - 06 July 1989
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Guideline study conducted according to GLP with acceptable restrictions: Limited details on test substance given.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 405 (Acute Eye Irritation / Corrosion)
- Deviations:
- yes
- Remarks:
- Limited details on study substance
- GLP compliance:
- yes
- Species:
- rabbit
- Strain:
- other: Kleinrusse, Chbb: HM
- Details on test animals or tissues and environmental conditions:
- TEST ANIMALS
- Source: Thomae, Biberach, Deutschland
- Age at study initiation: 6 months
- Weight at study initiation: 2550 g
- Diet: Altromin 2023, ad libitum
- Water: tap water, ad libitum
- Acclimation period: 1 week
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-25
- Humidity (%): 45-70
- 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:
- 24 h
- Observation period (in vivo):
- 72 h
Reading time points at 1, 24, 48 and 72 h - Number of animals or in vitro replicates:
- 3
- Details on study design:
- REMOVAL OF TEST SUBSTANCE
- Washing (if done): Yes, eye was rinsed with water
- Time after start of exposure: 24 h
SCORING SYSTEM: according to the Draize scoring system
TOOL USED TO ASSESS SCORE: fluorescein - Irritation parameter:
- cornea opacity score
- Basis:
- mean
- Remarks:
- out of all three 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 three animals
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 2
- Reversibility:
- other: reversibility not applicable
- Irritation parameter:
- conjunctivae score
- Basis:
- animal #1
- Time point:
- 24/48/72 h
- Score:
- 0.33
- Max. score:
- 3
- Reversibility:
- fully reversible within: 48 h
- Irritation parameter:
- conjunctivae score
- Basis:
- mean
- Remarks:
- out of animals #2 and #3
- 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 three animals
- Time point:
- 24/48/72 h
- Score:
- 0
- Max. score:
- 4
- Reversibility:
- other: reversibility not applicable
- Irritant / corrosive response data:
- None of the animals showed chemosis, corneal opacity and/or effects in the iris. In one case, conjunctival redness was noticed at 1 and 24 h, the 2 remaining animals did not show similar finding.
- Interpretation of results:
- not irritating
- Remarks:
- Migrated information Criteria used for interpretation of results: EU
- Conclusions:
- CLP: not classified
DSD: not classified - Endpoint:
- eye irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Guideline study with acceptable restrictions. No details on preparation of eyes before exposure. Application volume not reported. Limited details on test conditions and measurements.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)
- Deviations:
- no
- Principles of method if other than guideline:
- The test substance was investigated for the serious eye damaging potential using the standard BCOP protocol according to OECD guideline 437 and a modified surfactant protocol.
- GLP compliance:
- not specified
- Remarks:
- the limited information in the study report did not include a confirmation of the GLP status
- Species:
- other: cattle
- Details on test animals or tissues and environmental conditions:
- TEST METHOD
The bovine corneal opacity and permeability (BCOP) test is an in-vitro test method used to classify substances as “ocular corrosives and severe irritants”. The potential of a test substance to cause ocular corrosivity or severe irritancy is measured by its ability to induce opacity and increased permeability in an isolated bovine cornea. The opacity and permeability assessments of the cornea are combined to derive an in-vitro irritancy score (IVIS), which is used to classify the irritancy level of the test substance. However, the permeability value (OD490) is used as the primary endpoint for evaluation of the eye irritation potential for surfactant-based products.
IDENTIFICATION OF THE SOURCE OF THE EYES, STORAGE AND TRANSPORT CONDITIONS
- Donor animals: freshly slaughtered cattle
DETERMINATION OF THE INITIAL OPACITY
- Method: corneal opacity was determined by the amount of light transmission through the cornea via an opacitometer. Permeability is measured as the amount of sodium fluorescein dye that passes across the full thickness of the cornea. - Vehicle:
- other: undiluted or deionised water as vehicle
- Controls:
- other: number of eyes for the negative control: 3; number of eyes for the positive control: 3
- Amount / concentration applied:
- TEST MATERIAL
- Concentration (if solution): undiluted or 10% emulsion in deionised water (surfactant)
VEHICLE
- Substance: deionised water
POSITIVE SUBSTANCE
- Substance: sodium hydroxide
- Concentration (if solution): 1% - Duration of treatment / exposure:
- Standard protocol: 10 min (undiluted test substance)
Modified surfactant protocol: 1 h (undiluted test substance or 10% emulsion in deionised water) - Duration of post- treatment incubation (in vitro):
- Post-exposure incubation period: 2 h (standard protocol) or 1 h (modified surfactant protocol)
- Number of animals or in vitro replicates:
- number of eyes for the test item: 3
- Details on study design:
- TEST CONDITIONS
- Short description of the method used: the undiluted test substance or a 10% emulsion of the test substance in deionised water was topically applied to the epithelial surface of isolated corneas from the eyes of freshly slaughtered cattle. Control tissues were concurrently applied into the anterior chamber with de-ionised water (negative control, NC) or with sodium hydroxide in deionised water (positive control, PC). The corneas were incubated for 10 min.
DETERMINATION OF THE FINAL OPACITY
- Method: corneal opacity was determined by the amount of light transmission through the cornea via an opacitometer.
- Time of determination: directly after refilling fresh EMEM without phenol red in the anterior chamber, the final opacity was measured.
DETERMINATION OF THE CORNEAL PERMEABILITY:
- Method: permeability was measured via UV/VIS spectrophotometry at 490 nm recorded as optical density (OD490). - Irritation parameter:
- cornea opacity score
- Run / experiment:
- undiluted / 10 min
- Value:
- 5.3
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Irritation parameter:
- fluorescein leakage
- Run / experiment:
- undiluted/ 10 min
- Value:
- 0.002
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation parameter:
- in vitro irritation score
- Run / experiment:
- undiluted/ 10 min
- Value:
- 5.3
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Irritation parameter:
- cornea opacity score
- Run / experiment:
- undiluted/ 10 min
- Value:
- 96.9
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: positive control
- Irritation parameter:
- fluorescein leakage
- Run / experiment:
- undiluted/ 10 min
- Value:
- 3.463
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: positive control
- Irritation parameter:
- in vitro irritation score
- Run / experiment:
- undiluted/ 10 min
- Value:
- 148.9
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: positive control
- Irritation parameter:
- cornea opacity score
- Run / experiment:
- undiluted/ 10 min
- Value:
- 2.3
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: negative control
- Irritation parameter:
- fluorescein leakage
- Run / experiment:
- undiluted/ 10 min
- Value:
- -0.003
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: negative control
- Irritation parameter:
- in vitro irritation score
- Run / experiment:
- undiluted/ 10 min
- Value:
- 2.3
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: negative control
- Other effects / acceptance of results:
- Based on the measurements for opacity and permeability, an In Vitro Irritancy Score (IVIS) ≤ 55 was calculated after treatment with the test substance. Thus, no risk for serious damage to the eyes was identified. The positive and negative controls showed the expected results.
Using the standard BCOP protocol according to OECD guideline 437, no histopathological findings were observed in the corneas after treatment with the undiluted test substance. Using the modified surfactant protocol, histopathological examination revealed minimal eye damage in 2/3 corneas after exposure to the undiluted test substance. Treatment with 10% emulsion of the test substance did not reveal any histopathological findings indicative of eye damage using the modified surfactant protocol. - Interpretation of results:
- other: not corrosive
- Remarks:
- Criteria used for interpretation of results: EU
- Conclusions:
- There is regulatory acceptance in the EU that a substance can be considered as severe eye irritant (Serious eye damage Category 1/R41) based on a positive result in the Bovine Corneal Opacity and Permeability (BCOP) test. Negative in-vitro corrosivity responses are not conclusive with respect to non-classification or classification as irritant (Category 2/R36) and shall therefore be subject to further evaluation.
- Endpoint:
- eye irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: According to an accepted test protocol (EpiOcularTM Test; limited documentation)
- Principles of method if other than guideline:
- EpiOcularTM Test: The test substance is applicated to a reconstructed 3D human tissue sample for 30 min followed by a 2-hours post-incubation period. Tissue destruction is measured by MTT reduction.
- GLP compliance:
- not specified
- Remarks:
- the limited information in the study report did not include a confirmation of the GLP status
- Species:
- human
- Strain:
- other: EpiOcular™
- Details on test animals or tissues and environmental conditions:
- TEST SYSTEM
EpiOcular™ tissue sample (3-dimensional reconstructed human tissue) - Vehicle:
- unchanged (no vehicle)
- Controls:
- yes, concurrent positive control
- yes, concurrent negative control
- Amount / concentration applied:
- no data
- Duration of treatment / exposure:
- 30 min
- Number of animals or in vitro replicates:
- -Replicates: 2 tissue samples per condition
- Details on study design:
- Two EpiOcular™ tissue samples were incubated with the test substance for 30 minutes followed by a 2-hours post-incubation period.
Cell viability is measured by dehydrogenase conversion of the yellow, water-soluble MTT (3-[4,5-dimethylthazol-2-yl]-2,5-diphenyltetrazolium bromide) present in cell mitochondria, in to a blue formazan salt that is measured quantitatively after isopropanol extraction from the tissues. The optical density of the extracts of test substance treated tissues is compared to negative control values from tissue treated with PBS and is expressed as relative tissue viability. - Irritation parameter:
- mean percent tissue viability
- Run / experiment:
- 1
- Value:
- 100
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Other effects / acceptance of results:
- Mean cell viability: 100% (positive control: 19%)
- Interpretation of results:
- not irritating
- Conclusions:
- There is no regulatory acceptance in the EU that a substance can be considered "not irritating" based on a negative result in one of the available in vitro test methods, e.g. the EpiOcularTM. Further evaluation shoud be performed.
Referenceopen allclose all
Table 1: Eye irritation scores
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 |
|
1192 |
0 |
0 |
0 |
0 |
2 |
0 |
0 |
0 |
0 |
1 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
1193 |
0 |
0 |
0 |
0 |
1 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
1195 |
0 |
0 |
1 |
0 |
2 |
0 |
0 |
1 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
A = Corneal Score
B = Iris Score
C = Erythema Score
D = Chemosis Score
E = Exsudation Score
Table 1. Opacity scores after 10 min exposure and 2 h post-treatment incubation period
Treatment group |
Cornea no. |
Initial opacity |
Final opacity |
Opacity change |
Corrected opacity change |
Mean |
SD |
Test substance (undiluted) |
13 14 15 |
6.5 1.0 4.2 |
19.0 7.5 8.0 |
12.6 6.4 3.8 |
10.2 4.1 1.5 |
5.3 |
4.5 |
Negative control (deionised water) |
1 2 3 |
1.3 1.0 5.5 |
6.6 3.3 5.0 |
5.3 2.2 -0.5 |
NA NA NA |
2.3 |
2.9 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
2.9 1.3 4.3 |
90.6 99.1 116.7 |
87.7 97.7 112.4 |
85.3 95.4 110.1 |
96.9 |
12.4 |
SD = standard deviation; NA = not applicable
Table 2. Permeability score after 10 min exposure and 2 h post-treatment incubation period
Treatment group |
Cornea no. |
Mean OD490 |
Dilution factor |
Mean corrected OD490 |
Mean |
SD |
Test substance (undiluted) |
13 14 15 |
0.000 -0.001 -0.002 |
1 1 1 |
0.002 0.002 0.001 |
0.002 |
0.001 |
Negative control (deionised water) |
1 2 3 |
-0.009 0.001 0.000 |
1 1 1 |
NA NA NA |
-0.003 |
0.006 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
0.711 0.722 0.643 |
5 5 5 |
3.558 3.611 3.219 |
3.463 |
0.212 |
SD = standard deviation; NA = not applicable
Table 3. In vitro irritancy score (IVIS) after 10 min exposure and 2 h post-treatment incubation period
Treatment group |
Cornea no. |
Opacity per cornea |
Permeability per cornea |
IVIS |
||
per cornea |
per group |
|||||
mean |
SD |
|||||
Test substance (undiluted) |
13 14 15 |
10.2 4.1 1.5 |
0.002 0.002 0.001 |
10.3 4.1 1.5 |
5.3 |
4.5 |
Negative control (deionised water) |
1 2 3 |
5.3 2.2 -0.5 |
-0.009 0.001 0.000 |
5.2 2.2 -0.5 |
2.3 |
2.8 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
85.3 95.4 110.1 |
3.558 3.661 3.219 |
138.7 149.6 158.4 |
148.9 |
9.9 |
SD = standard deviation
Table 4. Opacity scores after 1 h exposure and 2 h post-treatment incubation period
Treatment group |
Cornea no. |
Initial opacity |
Final opacity |
Opacity change |
Corrected opacity change |
Mean |
SD |
First run (10% test substance preparation) |
|||||||
Test substance (10% in deionised water) |
10 11 12 |
2.7 3.8 3.5 |
2.8 3.4 3.5 |
0.1 -0.5 0.0 |
-0.1 -0.6 -0.2 |
-0.3 |
0.3 |
Negative control (deionised water) |
1 2 3 |
4.9 3.3 4.0 |
4.9 3.7 4.1 |
0.0 0.4 0.1 |
NA NA NA |
0.2 |
0.2 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
3.1 3.8 5.4 |
225.0 210.8 258.9 |
221.9 206.9 253.5 |
221.7 206.8 253.3 |
227.3 |
23.8 |
Second run (undiluted test substance) |
|||||||
Test substance (undiluted) |
7 8 9 |
6.3 3.4 5.1 |
7.7 3.0 5.5 |
1.4 -0.5 0.4 |
1.5 -0.4 0.5 |
0.6 |
0.9 |
Negative control (deionised water) |
1 2 3 |
4.3 2.4 3.7 |
3.6 2.5 4.0 |
-0.7 0.1 0.3 |
NA NA NA |
-0.1 |
0.5 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
1.5 2.3 5.1 |
228.0 213.5 255.1 |
226.5 211.2 250.0 |
226.7 211.3 250.1 |
229.4 |
19.6 |
SD = standard deviation; NA = not applicable
Table 5. Permeability score after 1 h exposure and 2 h post-treatment incubation period
Treatment group |
Cornea no. |
Mean OD490 |
Dilution factor |
Mean corrected OD490 |
Mean |
SD |
First run (10% test substance preparation) |
||||||
Test substance (10% in deionised water) |
10 11 12 |
0.000 0.000 -0.005 |
1 1 1 |
-0.014 -0.014 -0.019 |
-0.016 |
0.003 |
Negative control (deionised water) |
1 2 3 |
-0.007 -0.002 0.051 |
1 1 1 |
NA NA NA |
0.014 |
0.032 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
0.881 0.839 0.730 |
5 5 5 |
4.391 4.179 3.634 |
4.068 |
0.390 |
Second run (undiluted test substance) |
||||||
Test substance (undiluted) |
7 8 9 |
0.002 0.046 -0.003 |
1 1 1 |
0.005 0.049 0.000 |
0.018 |
0.027 |
Negative control (deionised water) |
1 2 3 |
-0.004 -0.003 -0.002 |
1 1 1 |
NA NA NA |
-0.003 |
0.001 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
0.639 0.820 0.608 |
5 5 5 |
3.200 4.102 3.045 |
3.449 |
0.571 |
SD = standard deviation; NA = not applicable
Table 6. In vitro irritancy score (IVIS) after 1 h exposure and 2 h post-treatment incubation period
Treatment group |
Cornea no. |
Opacity per cornea |
Permeability per cornea |
IVIS |
||
per cornea |
per group |
|||||
mean |
SD |
|||||
First run (10% test substance preparation) |
||||||
Test substance (10% in deionised water) |
10 11 12 |
-0.1 -0.6 -0.2 |
-0.014 -0.014 -0.019 |
-0.3 -0.8 -0.4 |
-0.5 |
0.3 |
Negative control (deionised water) |
1 2 3 |
0.0 0.4 0.1 |
-0.007 -0.002 0.051 |
-0.1 0.4 0.9 |
0.4 |
0.5 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
221.7 206.8 253.3 |
4.391 4.179 3.634 |
287.6 269.5 307.8 |
288.3 |
19.2 |
Second run (undiluted test substance) |
|
|||||
Test substance (undiluted) |
7 8 9 |
1.5 -0.4 0.5 |
0.005 0.049 0.000 |
1.6 0.4 0.5 |
0.8 |
0.6 |
Negative control (deionised water) |
1 2 3 |
-0.7 0.1 0.3 |
-0.004 -0.003 -0.002 |
-0.8 0.0 0.3 |
-0.2 |
0.6 |
Positive control (1% NaOH in deionised water) |
4 5 6 |
226.7 211.3 250.1 |
3.200 4.102 3.045 |
274.7 272.8 295.8 |
281.1 |
12.8 |
SD = standard deviation
Table 1: Results of the EpiOcularTM Test
Test substance |
|
Tissue 1 |
Tissue 2 |
Mean |
Inter tissue variability [%] |
NC |
Mean OD570 |
1.339 |
1.381 |
1.360 |
|
Viability [% of NC] |
98.4 |
101.6 |
100 |
3.1 |
|
Test item |
Mean OD570 |
1.409 |
1.314 |
1.361 |
|
Viability [% of NC] |
103.6 |
96.6 |
100 |
7.0 |
|
PC |
Mean OD570 |
0.257 |
0.263 |
0.260 |
|
Viability [% of NC] |
18.9 |
19.3 |
19 |
0.4 |
NC = negative control
PC = positive control
OD570 = Optical Density [wavelength 570 nm]
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (not irritating)
Respiratory irritation
Endpoint conclusion
- Endpoint conclusion:
- no study available
Additional information
Skin irritation
The skin irritation potential of Fatty acids, C18-unsatd., dimers, mixed esters with oleic acid and trimethylolpropane has been investigated in vivo and in vitro (CAS 147256-33-5).
Two studies in rabbits are available investigating the skin irritation potential of Fatty acids, C18-unsatd., dimers, mixed esters with oleic acid and trimethylolpropane according to OECD guideline 404 and in compliance with GLP (Pooles, 2012 and Steiling, 1989). In both studies, the clipped or shaved skin of three rabbits was exposed to 0.5 mL of the undiluted test substance for a period of 4 h under semi-occlusive conditions. The scoring of skin reactions (erythema and edema) was performed 1, 24, 48 and 72 h after removal of the dressing. Skin reactions involved the occurrence of erythema (grade 1) in 1/3 animals in both studies, which was fully reversible after 72 h (Oleon, 2012) and 7 days (BASF, 1989), respectively. No edema was observed in any of the treated animals. In one of the studies, the mean erythema and edema scores after 24, 48 and 72 h were 0/0.67/0 and 0 for the 3 animals (Oleon 2012). In the other study, the mean erythema and edema scores after 24, 48 and 72 h were 0/0/1 and 0 in the 3 animals (BASF, 1989).
In a further study, the skin corrosion/irritation potential of the test substance was investigated in an in vitro skin corrosion/irritation test using a human skin model according to OECD guidelines 431 and 439 (Remmele, 2012). In this GLP-compliant study, two replicates of intact reconstructed human epidermis (EpiDerm™) were topically treated with the undiluted test substance for 3 min or 1 h to assess the skin corrosion potential. The skin irritation test was performed in 3 replicate cultures treated for 1 h followed by a 42-h post-incubation period. Cells treated with de-ionized water (corrosion test) and PBS (irritation test) served as negative controls. After exposure to the test substance, cell viability was determined compared to the control. In the skin corrosion test, cell viability after 3 min exposure was greater than 50% (mean: 92%) and the cell viability after 1 h exposure was greater than 15% (mean: 100%). After 1-h treatment and 42-h post-exposure in the skin irritation test, cell viability was greater than 50% (mean: 112%). The positive controls 8 N KOH (corrosion test) and 5% SDS in de-ionized water (irritation test) showed the expected decrease in cell viability for the respective tests. Under the conditions of this in vitro study, the test material was non-corrosive to reconstructed human epidermis in vitro.
In summary, Fatty acids, C18-unsatd., dimers, mixed esters with oleic acid and trimethylolpropane is not irritating to the skin.
Eye irritation
CAS 147256-33-5
The eye irritation potential of Fatty acids, C18-unsatd., dimers, mixed esters with oleic acid and trimethylolpropane has been investigated in two studies in vivo and two studies in vitro (CAS 147256-33-5).
The eye irritation potential of Fatty acids, C18-unsatd., dimers, mixed esters with oleic acid and trimethylolpropane was investigated according to OECD guideline 405 and in compliance with GLP (Pooles, 2012). The undiluted test material (0.1 mL) was placed into the conjunctival sac of the right eye of 3 male New Zealand White rabbits. The other eye remained untreated and served as control. The eyes were examined and scored 1, 24, 48 and 72 h after application. Minimal conjunctival irritation (grade 1) was noted in all treated eyes 1 and 24 h after test substance instillation. However, these effects were fully reversible within 48 h. No corneal or iridial effects were noted at any observation time point during the study. No further local or systemic toxic effects were observed. The mean cornea, iris and chemosis scores after 24, 48, and 72 h were 0 for all 3 animals. The mean scores for conjunctivae after 24, 48 and 72 h were 0.33 for each individual animal. Based on these results, the test substance is not considered to be an eye irritant.
A further eye irritation study with the test substance was performed in Kleinrussen, Chbb: HM rabbits according to OECD guideline 405 and under conditions of GLP (Steiling, 1989). The undiluted test substance (0.1 mL) was placed into one eye of 3 male Small White Russian (Mol. Russian, Chbb:MH) rabbits for a period of 24 h and then rinsed with water. The other eye remained untreated and served as control. Examination and scoring of effects on the eyes was performed at intervals of 1, 24, 48 and 72 h after test substance application. None of the treated animals showed chemosis, corneal opacity and/or effects on the iris. In a single animal, slight conjunctival redness was noticed at 1 and 24 h, which was fully reversible at the 72 h reading. No further local or systemic toxic effects were observed. The mean cornea, iris and chemosis scores after 24, 48, and 72 h were 0 for all 3 animals, respectively. The mean scores for conjunctivae after 24, 48, and 72 h were 0.33/0/0 for the 3 individual animals. Based on these results, the test substance is not considered to be an eye irritant.
The non-irritating potential of the test substance was further supported in a GLP-compliant in vitro eye irritation test using the reconstructed human cornea model EpiOcularTM (Remmele, 2012). In this experiment, two samples of tissue were incubated with the test substance for 30 min followed by a 2-h post-incubation period. Cells treated with sterile de-ionised water served as controls. After treatment, the mean cell viability compared to control was determined to be 100%. Thus, cell survival was not influenced by treatment with the test substance. The positive control methyl acetate fulfilled the criteria for a positive result (< 50% mean cell viability). Under the conditions of this study, the test substance was not irritating to reconstructed human cornea in vitro.
In a Bovine Corneal Opacity and Permeability (BCOP) test, the potential for corrosivity and severe irritation was investigated in vitro according to the standard protocol as described in OECD guideline 437 and according to a modified surfactant protocol (Remmele, 2012). Using the guideline standard protocol, the undiluted test substance was applied to the epithelial surface of the isolated cornea of cattles for 10 min, followed by a 2-h post-exposure incubation period. According to a modified surfactant protocol, the isolated corneas of cattles were exposed for 1 h to the undiluted test substance or a 10% emulsion in deionised water, followed by a 2-h exposure period. After exposure, no changes in opacity and permeability were observed in corneas compared to control, neither using the standard nor the modified test protocol. Based on the measurements for opacity and permeability, an In Vitro Irritancy Score (IVIS) ≤ 55 was calculated after treatment with the test substance using both test protocols. Histological evaluation was performed in addition to the assessment of opacity and permeability. Using the standard BCOP protocol according to OECD guideline 437, no histopathological findings were observed in the corneas after treatment with the undiluted test substance. Using the modified surfactant protocol, histopathological examination revealed minimal eye damage in 2/3 corneas after exposure to the undiluted test substance. Treatment with 10% emulsion of the test substance did not reveal any histopathological findings indicative of eye damage using the modified surfactant protocol. The positive (1% sodium hydroxide) and negative (deionised water) controls showed the expected results and thus verified the reliability of the assay. Based on the results of this study, no risk for serious damage to the eyes was identified.
In summary, Fatty acids, C18-unsatd., dimers, mixed esters with oleic acid and trimethylolpropane is not eye irritating.
Conclusions for irritation / corrosion
The substance Fatty acids, C18-unsatd., dimers, mixed esters with oleic acid and trimethylolpropane has been tested for skin and eye irritation in vivo and in vitro, showing that the substance is not irritating to the skin and eyes.
Justification for classification or non-classification
The available data on skin and eye irritation of Fatty acids, C18-unsatd., dimers, mixed esters with oleic acid and trimethylolpropane do not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.
