Registration Dossier

Administrative data

Description of key information

The in vitro skin corrosion test was conducted according to OECD 431 guideline and GLP principles. The test substance is not corrosive in the in vitro skin corrosion test.
In a skin irritation study with rabbits, performed according to OECD 404 guideline and GLP principles, very slight erythema was observed. The test substance is considered to be not irritating to skin.
The test substance is not irritant or corrosive in the Bovine Corneal Opacity and Permeability test performed according to OECD 437 guideline and GLP principles.
In an eye irritation study with rabbits, performed according to OECD 405 guideline and GLP principles, irritation was observed. The test substance is considered to be not irritating to eyes.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records
Reference
Endpoint:
skin irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
26 March - 05 April 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2500 (Acute Dermal Irritation)
Deviations:
no
Qualifier:
according to
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries (JMAFF), 12 Nohsan, Notification No 8147, April 2011, including the most recent partial revisions.
Deviations:
no
GLP compliance:
yes (incl. certificate)
Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
- Source: Charles River Deutschland, Kisslegg, Germany
- Age at study initiation: Animals used withihn the study were at least 6 weeks old.
- Weight at study initiation: Body weights were at least 1.0 kg .
- Housing: Animals were housed individually in cages with perforated floors.
- Diet: Free access to pelleted diet for rabbits (Global Diet 2030 from Harlan Teklad, Italy). Hay and wooden sticks were available during the study period.
- Water: Free access to tap water.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, approximately 15 room air changes/hour, and a 12-hour light/12-hour dark cycle. Any variations to these conditions were maintained in the raw data and had no effect on the outcome of the study.

IN-LIFE DATES: From: 26 March - 05 April 2013
Type of coverage:
semiocclusive
Preparation of test site:
shaved
Vehicle:
unchanged (no vehicle)
Controls:
other: Adjacent areas of the untreated skin of each animal served as controls.
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 mL
Duration of treatment / exposure:
Single application.
Observation period:
14 days.
Number of animals:
3 males.
Details on study design:
STUDY DESIGN
The study was performed in a stepwise manner and was started by treatment of a single rabbit (sentinel). The two other animals were treated in a similar manner 7 days later, after considering the degree of skin irritation observed in the first animal.

TEST SUBSTANCE PREPARATION
The test substance was applied undiluted as delivered by the sponsor. No corrections were made for the purity/composition of the test substance, since the guidelines requires a fixed amount that has to be applied.

TEST SITE
Approximately 24 hours before treatment, the dorsal fur was clipped with electric clippers, exposing an area of approximately 150 square centimeters (10x15 cm). To facilitate scoring, treated skin areas were re-clipped at least 3 hours before the observations.

The test substance was applied to the skin of one flank, using a patch of 2x3 cm.

REMOVAL OF TEST SUBSTANCE
Four hours after the application, the dressing was removed and the skin cleaned of residual test substance using tap water and watery ethanol (50% v/v).

OBSERVATIONS
- Mortality/Viability: Twice daily.
- Toxicity: At least once daily.
- Body Weight: Day of treatment (prior to application) and after the final observation.
- Necropsy: No necropsy was performed according to protocol.
- Irritation: The skin reactions were assessed at approximately 1, 24, 48 and 72 hours after the removal of the dressings and test substance. The irritation scores and a description of all other (local) effects were recorded. Adjacent areas of the untreated skin of each animal served as controls.

SCORING SYSTEM:
The irritation was assessed according to the numerical scoring system according to OECD 404.
Irritation parameter:
erythema score
Basis:
mean
Time point:
24/48/72 h
Score:
0.2
Max. score:
4
Reversibility:
fully reversible within: 48 hours after exposure in two animals and within 24 hours in the remaining animal.
Irritation parameter:
edema score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
4
Irritant / corrosive response data:
Irritation: Four hours exposure to 0.5 mL of m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline resulted in very slight erythema and very slight oedema in the treated skin areas of the three rabbits. The skin irritation resolved within 48 hours after exposure in two animals and within 24 hours in the remaining animal.
Corrosion: There was no evidence of a corrosive effect on the skin.
Other effects:
Coloration/remnants: No staining of the treated skin by the test substance was observed. Dry remnants of the test substance were present on the skin on Day 1.
Toxicity/mortality: No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
In a skin irritation study with rabbits, performed according to OECD 404 guideline and GLP principles, very slight erythema was observed. The skin irritation was fully reversible and therefore m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline does not have to be classified and has no obligatory labelling requirement.
Executive summary:

m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline was tested in a skin irritation study with rabbits, performed according to OECD 404 guideline and GLP principles.

Four hours exposure to m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline resulted in very slight erythema and very slight oedema in the treated skin areas of the three rabbits. The skin irritation resolved within 48 hours after exposure in two animals and within 24 hours in the remaining animal. There was no evidence of a corrosive effect on the skin.

No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.

Based on the results, m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline does not have to be classified and has no obligatory labelling requirement for skin irritation according Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
02 - 16 April 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2400 (Acute Eye Irritation)
Deviations:
no
Qualifier:
according to
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries (JMAFF), 12 Nohsan, Notification No 8147, April 2011, including the most recent partial revisions.
Deviations:
no
GLP compliance:
yes (incl. certificate)
Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
- Source: Charles River France, L’Arbresle Cedex, France
- Age at study initiation: Animals used withihn the study were at least 6 weeks old.
- Weight at study initiation: Body weights were at least 1.0 kg.
- Housing: Individually housed in cages with perforated floors.
- Diet: Free access to pelleted diet for rabbits (Global Diet 2030 Harlan Teklad, Italy). Hay and wooden sticks were available during the study period.
- Water: Free access to tap water.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, approximately 15 room air changes/hour, and a 12-hour light/12-hour dark cycle. Any variations to these conditions were maintained in the raw data and had no effect on the outcome of the study.

IN-LIFE DATES: From: 02 - 16 April 2013
Vehicle:
unchanged (no vehicle)
Controls:
other: One eye of each animal remained untreated and served as the reference control.
Amount / concentration applied:
TEST MATERIAL- Amount applied: 0.1 mL
Duration of treatment / exposure:
Single instillation on Day 1.
Observation period (in vivo):
The eyes of each animal were examined approximately 1, 24, 48 and 72 hours and 7 days after instillation of the test substance. The irritation scores and a description of all other (local) effects were recorded.
Number of animals or in vitro replicates:
3 males
Details on study design:
STUDY DESIGN
The study was performed in a stepwise manner and was started by treatment of a single rabbit (sentinel). The two other animals were treated in a similar manner one week later, after considering the degree of eye irritation observed in the first animal.

PREEMPTIVE PAIN MANAGEMENT
Both eyes were topically anaesthetized using a local anaesthetic (Lidocaine eye drops, AST Farma BV, Woerden, The Netherlands). Approximately 10 minutes prior to treatment, 1 drop was applied to the eyes. No routine pretreatment with a systemic analgesic was done. The subcutaneous injection was
considered to cause unwanted additional distress, since in the majority of cases instillation of test materials does not cause much distress in the topically anesthetized eye. No post-treatment with systemic analgesics were needed.

TREATMENT
Each animal was treated by instillation of 0.1 mL of the test substance, in the conjunctival sac of one of the eyes after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about one second to prevent loss of the test substance. The other eye remained untreated and served as the reference control. Immediately after the 24-hour observation, a solution of 2% fluorescein (Merck, Darmstadt, Germany) in water (adjusted to pH 7.0) was instilled into both eyes of each animal to quantitatively determine corneal epithelial damage. This procedure was repeated to assess recovery in two animals. Any bright green stained area, indicating epithelial damage, was estimated as a percentage of the total corneal area.
After the final observation, the animals were sacrificed by intra-venous injection of Euthasol® 20% (AST Farma BV, Oudewater, The Netherlands).

REMOVAL OF TEST SUBSTANCE
-Washing: No

OBSERVATIONS
- Mortality/Viability: Twice daily.
- Toxicity: At least once daily.
- Body Weight: Day of treatment (prior to instillation) and after the final observation.
- Necropsy: No necropsy was performed according to protocol.
- Irritation: The eyes of each animal were examined approximately 1, 24, 48 and 72 hours and 7 days after instillation of the test substance. The irritation scores and a description of all other (local) effects were recorded.
Irritation parameter:
cornea opacity score
Remarks:
(opacity)
Basis:
mean
Remarks:
(3 animals)
Time point:
24/48/72 h
Score:
0
Max. score:
4
Irritation parameter:
iris score
Basis:
mean
Remarks:
(3 animals)
Time point:
24/48/72 h
Score:
0.3
Max. score:
2
Reversibility:
fully reversible within: 48 or 72 hours
Irritation parameter:
conjunctivae score
Remarks:
(redness)
Basis:
mean
Remarks:
(3 animals)
Time point:
24/48/72 h
Score:
1.6
Max. score:
3
Reversibility:
fully reversible within: 7 days
Irritation parameter:
chemosis score
Basis:
mean
Remarks:
(3 animals)
Time point:
24/48/72 h
Score:
1.2
Max. score:
4
Reversibility:
fully reversible within: 7 days
Irritant / corrosive response data:
Irritation: Instillation of 0.1 mL of m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline into one eye of each of three rabbits resulted in effects on the cornea, iris and conjunctivae. Slight dulling of the normal luster of the cornea was observed after 24 hours in two animals (maximum 25% of the corneal area) and after 48 hours in the remaining animal (maximum 50% of the corneal area). Fluorescein staining showed epithelial damage on the cornea (maximum 10% of the corneal area) in two animals. The corneal injury resolved within 72 hours. Iridial irritation grade 1 was observed in two animals and resolved within 48 or 72 hours. The irritation of the conjunctivae consisted of redness, chemosis and discharge and completely
resolved within 7 days in all animals.
Corrosion: There was no evidence of ocular corrosion.
Other effects:
Colouration/Remnants: No staining of (peri) ocular tissues by the test substance was observed. Remnants of the test substance were present on the outside of the eyelids of all animals on Days 1 and 2.
Toxicity/Mortality: No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.
Interpretation of results:
other: \\\Slightly irritating to eyes, but insufficient for classification.
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
In an eye irritation study with rabbits, performed according to OECD 405 guideline and GLP principles, irritation was observed.
Based on the results m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline does not have to be classified and has no obligatory labelling requirement for skin irritation according.
Executive summary:

m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline was tested in an eye irritation study with rabbits, performed according to OECD 405 guideline and GLP principles.

Instillation of m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline into one eye of each of three rabbits resulted in effects on the cornea, iris and conjunctivae. Slight dulling of the normal luster of the cornea was observed after 24 hours in two animals (maximum 25% of the corneal area) and after 48 hours in the remaining animal (maximum 50% of the corneal area). Fluorescein staining showed epithelial damage on the cornea (maximum 10% of the corneal area) in two animals. The corneal injury resolved within 72 hours. Iridial irritation grade 1 was observed in two animals and resolved within 48 or 72 hours. The irritation of the conjunctivae consisted of redness, chemosis and discharge and completely resolved within 7 days in all animals. There was no evidence of ocular corrosion.

No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.

Based on the results m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline does not have to be classified and has no obligatory labelling requirement for eye irritation according to Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

In vitro skin corrosion test:

In an in vitro skin corrosion test using a human skin model (EpiDerm Skin Model) performed according to OECD 431 guideline and GLP principles, the influence of the test substance on the viability of human skin was tested.

The test substance was applied directly to 0.6 cm^2 cultured skin (25 mg, in presence of 25 ul Milli-Q water). After 3 minutes and 1 hour, the substance was removed and cells were cultured for 42 hours. The viability of the cells was tested by reduction of MTT. The test substance did interact with MTT. In addition to the normal 1-hour procedure, one freeze-killed tissue treated with test substance and one freeze-killed non treated tissue was used for the cytotoxicity evaluation with MTT. The non-specific reduction of MTT by the test substance was 0.47% of the negative control tissues. The net OD of the treated freeze-killed tissues was subtracted from the ODs of the test substance treated viable tissues.

Survival of unexposed skin was set at 100%, the positive control had a mean cell viability of 10% after 3 minutes exposure whereas the test substance showed cell viability of 117% and 96% after 3-minute and 1-hour treatments, respectively. Because the mean relative tissue viability for the test substance was not below 50% after the 3-minute treatment and not below 15% after the 1-hour treatment, the test substance is considered to be not corrosive.

In vivo skin irritation test:

m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline was tested in a skin irritation study with rabbits, performed according to OECD 404 guideline and GLP principles.

Four hours exposure to m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline resulted in very slight erythema and very slight oedema in the treated skin areas of the three rabbits. The skin irritation resolved within 48 hours after exposure in two animals and within 24 hours in the remaining animal. There was no evidence of a corrosive effect on the skin.

No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred. The test substance is considered to be not irritating to skin.

In vitro eye irritation test:

A Bovine Corneal Opacity and Permeability test was performed with the test substance according to OECD 437 guideline and GLP principles.

Approx. 750 µl test substance was applied, in total 3 cornea's were tested with the test substance. The mean in vitro irritancy score was 1.0 after 10 minutes of treatment with the test substance. Since the mean in vitro irritancy score for the test substance was below 55.1 after 10 minutes treatment, the test substance is considered to be not severely irritant or corrosive.

In vivo eye irritation test:

m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline was tested in an eye irritation study with rabbits, performed according to OECD 405 guideline and GLP principles.

Instillation of m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline into one eye of each of three rabbits resulted in effects on the cornea, iris and conjunctivae. Slight dulling of the normal luster of the cornea was observed after 24 hours in two animals (maximum 25% of the corneal area) and after 48 hours in the remaining animal (maximum 50% of the corneal area). Fluorescein staining showed epithelial damage on the cornea (maximum 10% of the corneal area) in two animals. The corneal injury resolved within 72 hours. Iridial irritation grade 1 was observed in two animals and resolved within 48 or 72 hours. The irritation of the conjunctivae consisted of redness, chemosis and discharge and completely resolved within 7 days in all animals. There was no evidence of ocular corrosion.

No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred. The test substance is considered to be not irritating to eyes.


Justification for selection of skin irritation / corrosion endpoint:
One in vivo study available with klimisch score 1.

Justification for selection of eye irritation endpoint:
One in vivo study available with klimisch score 1.

Justification for classification or non-classification

Based on the above results m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline does not have to be classified and has no obligatory labelling requirement for skin and eye irritation according to Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures.