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Description of key information

Two valid and reliable studies were performed with commercial products following the principles of GLP. Pigment Red 57:1 did not cause skin sensitization in the Buehler test in guinea pigs (ETAD 1995) and in the local lymph node assay in mice (Clariant 2008).

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:
January - March 2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of study:
mouse local lymph node assay (LLNA)
Specific details on test material used for the study:
- Name of test material (as cited in study report): Graphtol-Rubine 6BP
- Substance type: pigment
- Physical state: red solid
- Expiration date of the lot/batch: October 31, 2017
- Stability under test conditions: stable
- Storage condition of test material: room temperature
- Other: stability > 72 hours in DMSO and 1,2 - propylene glycol at room temperature
- Analytical purity: 84.34/ (w/w)
- Composition: 7.73 % Pigment Red 63:1
- Lot/batch No.: KRON 792014
Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Netherlands
- Age at study initiation: 8 - 10 weeks
- Housing: single caging
- Diet (e.g. ad libitum): pelleted standard diet, ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 + 3°C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12h/12h
Vehicle:
dimethyl sulphoxide
Concentration:
2.5, 5, and 10 %
No. of animals per dose:
4
Details on study design:
For determination of the highest non-irritant and technically applicable test item concentration, a non-GLP pretest was performed on two mice with concentrations of 1.25, 2.5, 5, and 10 % (w/v). The top dose of the test item is the highest technically achievable concentration whilst avoiding systemic toxicity and excessive local irritation. Four female mice were treated with different concentrations of the test item and vehicle alone by topical application at the dorsum of each ear lobe (left and right) on three consecutive days. Five days after the first topical application, the mice were intravenously injected into a tail vein with radio-labelled thymidine (3H-methyl thymidine). Five hours after intravenous injection, the mice were sacrificed and the draining auricular lymph nodes excised and pooled per group. Single cell suspensions of lymph node cells were prepared from pooled lymph nodes which were subsequently washed and incubated with trichloroacetic acid overnight. The proliferative capacity of pooled lymph node cells was determined by the incorporation of 3H-methyl thymidine measured in a ß-scintillation counter.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Positive control results:
Performed in October 2007: 25% in acetone/olive oil (4+1) gave an SI of 6.32 and 10% gave an SI of 3.03.
Parameter:
SI
Value:
2
Test group / Remarks:
25%
Parameter:
SI
Value:
2.06
Test group / Remarks:
10%
Parameter:
SI
Value:
1.68
Test group / Remarks:
5%
Parameter:
SI
Value:
1
Test group / Remarks:
vehicle

Disintegrations per minute

DPM minus background control: 10594 (25%) 10939 (10%) 8907 (5%) 5298 (vehicle control)

Other information

No deaths occurred during the study period. Due to the intense red colour of the test item local irritation reactions such as ear redness could not be detected.

The body weights of the animals, recorded prior to the 1st application and prior to necropsy was within the range commonly recorded for animals of this strain and age.

Interpretation of results:
GHS criteria not met
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

The first study (ETAD 1995) is a test in guinea pigs (Buehler, 1965, described in OECD testing guideline 406, adopted July 17, 1992) and was performed with a sample of adequately high pigment content. Due to the staining properties of the pigment, skin reactions could not be examined visually and were investigated by histopathology (acanthosis, inflammatory cell infiltration, spongiosis) instead. The highest concentration of 60% for both challenge and induction was chosen based on the solubility in the vehicle corn oil. A predominantly slight to minimal inflammatory reaction was recorded in about half of the animals of both control and test groups. Therefore, Pigment Red 57:1 was evaluated as non-sensitizing in the Buehler test.

The second study (Clariant 2008) is a local lymph node assay in mice (OECD 429, adopted in April 24, 2002) and the tested sample is also of adequately high pigment content. Tested concentrations were 2.5, 6 and 10% using DMSO as vehicle and resulting in stimulation indices of 1.68, 2.06 and 2.00, respectively. The highest concentration was justified by the limit of solubility in DMSO. Higher concentrations could be achieved in DMF, but the sample was demonstrated to be unstable in this solvent. Due to the intense red colour of the test item local irritation reactions such as ear redness could not be evaluated. Based on the stimulation indices, the pigment is considered not to be a skin sensitizer in mice. 

Both studies are adequate to assess the endpoint of skin sensitization. The study by Clariant (2008) is chosen as key study because more detailed information in regard to sample composition and choice of vehicle.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. As a result the substance is not considered to be classified for skin sensitization according to Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008.