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Description of key information

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
October 31, 2012 - November 19, 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
(2010)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
(2008)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Version / remarks:
(2003)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
other: CBA/J
Sex:
female
Details on test animals and environmental conditions:
- Source: Janvier, Le Genest-Saint-Isle, France
- Age at study initiation: Young adult animals (approx. 10 weeks old)
- Weight at study initiation: Body weight variation was within +/- 20% of the sex mean.
- Housing: Animals were group housed in labeld makrolon cages.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap water.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 – 24
- Humidity (%): 40 - 70
- Air changes (per hr): approx 15
- Photoperiod (hrs dark / hrs light): 12/12
Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
0, 1, 2.5 and 5%
No. of animals per dose:
5
Details on study design:
RANGE FINDING TESTS:
Initially, two test substance concentrations were tested; a 25% and 50% concentration. The highest concentration was the maximum concentration as required in the test guidelines (50% for solids). The test system, procedures and techniques were identical to those used in the main study except that assessment of lymph node proliferation and necropsy were not performed. Two young adult animals per concentration were selected (in the range of 8 to14 weeks old). Each animal was treated with one concentration on three consecutive days.Based on the results of the initially treated animals, four additional animals were treated in a similar manner with two lower concentrations (5% and 10%) at a later stage.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response: DPM values are presented for each animal and for each dose group. A Stimulation Index (SI) is calculated for each group. The SI is the ratio of the DPM/group compared to DPM/vehicle control group. If the results indicate a SI ≥ 3, the test substance may be regarded as a skin sensitizer.

ANIMAL ASSIGNMENT
Three groups of five animals were treated with one test substance concentration per group. One group of five animals was treated with vehicle.

TREATMENT PREPARATION AND ADMINISTRATION:
Test substance preparation: The test substance formulations (w/w) were prepared within 4 hours prior to each treatment. No adjustment was made for specific gravity of the vehicle. Homogeneity was obtained to visually acceptable levels.
Correction of the purity/composition of the test substance is not applicable, since the test method requires a logical concentration range rather than specific dose levels to be applied.
Rationale for vehicle: The vehicle was selected based on trial formulations performed at WIL Research Europe and on test substance data supplied by the sponsor.

Induction - Days 1, 2 and 3; Excision of nodes - Day 6; Tissue processing for radioacitivity - Day 6; Radioactivity measurements - Day 7; Performed according to test guidelines.

Observations:
Mortality/Viability: Twice daily.
Body weights: On Day 1 (pre-dose) and Day 6 (prior to necropsy).
Clinical signs: Once daily on Days 1-6 (on Days 1-3 between 3 and 4 hours after dosing).
Irritation: Once daily on Days 1-6 (on Days 1 - 3 within 1 hour after dosing) according to the numerical scoring system. Furthermore, a description of all other (local) effects was recorded according to guidelines.

Necropsy: No necropsy was performed according to protocol.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
Not performed.
Positive control results:
The six-month reliability check with Alpha-hexylcinnamicaldehyde indicates that the Local Lymph Node Assay as performed at WIL Research Europe is an appropriate model for testing for contact hypersensitivity. See attached document 'Reliability check'.
Key result
Parameter:
SI
Value:
1.4
Variability:
1% concentration
Test group / Remarks:
5
Remarks on result:
other: The SI values calculated for the substance concentrations 1, 2.5 and 5% were 1.4, 4.3 and 12.1 respectively.
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: Mean DPM/animal values for the experimental groups treated with test substance concentrations 1, 2.5 and 5% were 721, 2238 and 6299 DPM respectively. The mean DPM/animal value for the vehicle control group was 519 DPM.
Key result
Parameter:
SI
Value:
4.3
Variability:
2.5% concentration
Test group / Remarks:
5
Remarks on result:
other: The SI values calculated for the substance concentrations 1,2.5 and 5% were 1.4, 4.3 and 12.1 respectively.
Key result
Parameter:
SI
Value:
12.1
Variability:
5% concentration
Test group / Remarks:
5
Remarks on result:
other: The SI values calculated for the substance concentrations 1, 2.5 and 5% were 1.4, 4.3 and 12.1 respectively.

Results Pre-screen test:

At a 5% test substance concentration no signs of systemic toxicity were noted and only very slight erythema was observed. At this dose level of 5% the variations in ear thickness during the observation period were less than 25% from Day 1 pre-dose values.

At higher concentrations (10, 25 and 50%) increasing irritation was noted and the variations in ear thickness during the observation period were above 25% from Day 1 pre-dose values.

Based on these results, the highest test substance concentration selected for the main study was a 5% concentration.

Results - main study:

The very slight irritation of the ears as shown by the animals treated at 5% was considered not to have a toxicologically significant effect on the activity of the nodes. No irritation was noted in the other animals.

 

The auricular lymph nodes of all animals at 5% were considered larger in size compared to control group. All auricular lymph nodes of the other animals of the experimental and control groups were considered normal in size.

No macroscopic abnormalities of the surrounding area were noted in any of the animals.

 

No mortality occurred and no clinical signs of systemic toxicity were observed in the animals of the main study. Body weights and body weight gain of experimental animals remained in the same range as controls over the study period. The slight body weight changes noted for some animals across the dose groups was considered not toxicologically significant since the changes were slight in nature and no concentration-related incidence was apparent. Only one animal at 5% (an. no. 18) showed moderate body weight loss (4g), and a lean appearance was observed on Day 6. This incidental finding was considered not toxicologically significant based on the absence of corroborative findings in this animal.   

Interpretation of results:
sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
In an LLNA skin sensitisation study, performed according to OECD 429 test guideline and GLP principles, m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline was considered to be a skin sensitiser, as the SI appeared to be ≥ 3 when tested up to 5%.
Executive summary:

The test substance was assessed for contact hypersensitivity in an LLNA skin sensitisation study up to 5%, performed according to OECD 429 test guideline and GLP principles.

The auricular lymph nodes of all animals at 5% were considered larger in size compared to control group. All auricular lymph nodes of the other animals of the experimental and control groups were considered normal in size.

No macroscopic abnormalities of the surrounding area were noted in any of the animals.

No mortality occurred and no clinical signs of systemic toxicity were observed in the animals of the main study.

The SI values calculated for the substance concentrations 1, 2.5 and 5% were 1.4, 4.3 and 12.1 respectively. These results indicate that the test substance could elicit an SI ≥ 3 when tested up to 5%.

Based on these results, the test substance should be classified as skin sensitizer (Category 1A) and labeled as H317: May cause an allergic skin reaction according to Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)
Additional information:

The test substance was assessed for contact hypersensitivity in an LLNA skin sensitisation study up to 5%, performed according to OECD 429 test guideline and GLP principles.

The auricular lymph nodes of all animals at 5% were considered larger in size compared to control group. All auricular lymph nodes of the other animals of the experimental and control groups were considered normal in size.

No macroscopic abnormalities of the surrounding area were noted in any of the animals.

No mortality occurred and no clinical signs of systemic toxicity were observed in the animals of the main study.

The SI values calculated for the substance concentrations 1, 2.5 and 5% were 1.4, 4.3 and 12.1 respectively. These results indicate that the test substance could elicit an SI ≥ 3 when tested up to 5% and is considered to be a skin sensitiser.


Migrated from Short description of key information:
In an LLNA skin sensitisation study, performed according to OECD 429 test guideline and GLP principles, m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline was considered to be a skin sensitiser, as the SI appeared to be ≥ 3 when tested up to 5%.

Justification for selection of skin sensitisation endpoint:
One study with klimisch code 1 is available.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Based on the results, m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline should be classified as skin sensitizer (Category 1A) and labeled as H317: May cause an allergic skin reaction according to Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures.