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Description of key information

Not sensitising to skin as assessed in guinea pig maximization assays (OECD 406) and local lymph node assay (OECD 429) with 'yellow diazo condensation pigments' (Read-across to CAS 68516-734 and CAS 5580-57-4).

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
READ ACROSS ANALOGUE APPROACH

The read-across hypothesis is fundamentally based on the same core structure of five ‘yellow disazo condensation pigments’ which optionally can serve as target or as source substances. None of the pigments are sufficiently soluble, either in water or in octanol for systemic uptake or metabolism. The molecular weight ranges from 716.6 g/mol (Pigment Yellow 155) to 1229.2 g/mol (Pigment Yellow 128). Therefore, the molecular weights of all ‘yellow disazo condensation pigments’ are well above the threshold of 500 g/mol, which is generally considered for low dermal and oral uptake [ECHA Guidance R. 7c, 2017]. Furthermore, for each of the substances, the critical body burden (CBB) is above the octanol solubility, which generally indicates a low uptake in biota and makes toxicity unlikely [ECHA Guidance R. 11, 2017].


Please find the complete Read-across justification text including a data matrix and structures attached in the attachment.

Reason / purpose for cross-reference:
read-across source
Key result
Parameter:
other: LN cell count index
Value:
0.84
Test group / Remarks:
0.1% test substance
Key result
Parameter:
other: LN cell count index
Value:
0.84
Test group / Remarks:
1% test substance
Key result
Parameter:
other: LN cell count index
Value:
1.04
Test group / Remarks:
10% test substance
Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
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)
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)
Specific details on test material used for the study:
- Name: Pigment Yellow 93
-appearance: yellow powder
-purity: 98%
- Lot/batch No.:7218
- Expiration date of the lot/batch: 02/ 2023
- Stability under test conditions: stable
- Storage condition of test material: The substance was indicated to be stored in the airtight delivered plastic container in the darkness at room temperature
Species:
mouse
Strain:
Balb/c
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Breeding farm VELAZ s.r.o., Kolec u Kladna, Czech Republic, ReH CZ 21760118
- Age at study initiation: 8 to 10 weeks (at start of dosing)
- Weight at study initiation: 17,3 to 21,6 g (at start of dosing)
- Housing: in groups of maximum six
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 hours light/dark cycle

IN-LIFE DATES: From: 3.11.2008 To: 5.11.2008
Vehicle:
other: DAE 433 - mixture of40% dimethylacetamide, 30% acetone and 30% ethanol
Concentration:
0.1, 1 and 10%
No. of animals per dose:
6
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: The test item is a pigment and insoluble in organic or inorganic solvents. A suspension of up to 10% could be tested.
- Irritation: Not irritating
- Lymph node proliferation response: No data

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: non-radiolabelled LLNA
- Criteria used to consider a positive response: The results of the LLNA were evaluated according to the following criteria.
The following thresholds were determined by Ulrich (2007) from analysis of historical data:
Ear weight index: 1.05
LN weight: 1.2
LN cell count: 1.3

1. Values which exceed these thresholds were considered positive
- when a statistically significant increase in one of the parameters occurs and a clear
concentration-dependence can be derived
- or with no statistical significance, but a clear concentration-dependence.
2. Values which are below these thresholds were considered positive
- when a statistical significant occurs in one of the parameters together with a clear
concentration dependence.

TREATMENT PREPARATION AND ADMINISTRATION: Before start of application the suspension was mixed for 5 minutes with a magnetic stirrer and then were still mixed during application.
Positive control substance(s):
other: Dinitrochlorobenzene
Statistics:
For statistical calculations the software Statgraphic® Centurion (version XV, USA) was used. At first the global comparison of all three values of the concentration groups with vehicle control was performed by applying the non-parametric Kruskal-Wallis test, and then the non-parametric two-group Mann-Whitney rank test (probability level 0.05) was applied to all two-group comparisons.
Parameter:
other: LN cell count index
Value:
1
Test group / Remarks:
Negative control
Parameter:
other: LN cell count index
Value:
4.06
Test group / Remarks:
Positive control
Key result
Parameter:
other: LN cell count index
Value:
0.84
Test group / Remarks:
0.1% test substance
Key result
Parameter:
other: LN cell count index
Value:
0.84
Test group / Remarks:
1% test substance
Key result
Parameter:
other: LN cell count index
Value:
1.04
Test group / Remarks:
10% test substance

Mean local lymph node weight (mg)

positive control: 14,87

Negative contro: 5,68

Test substance 0.1% : 5,77

Test substance 1%: 5,68

Test substance 10%: 5,45

Interpretation of results:
GHS criteria not met
Conclusions:
At the given experimental conditions the test substance elicited negative result in LLNA test. Based on these results the test substance does not have to be classified as the substance, which may cause sensitisation by skin contact.
Executive summary:

In this study the contact allergenic potential of the test item was evaluated with non radioactive measuring of cell proliferation after topical application to female BALB/c mice. Six mice per group were exposed by test and control substances on the dorsum of both ears once a day during 3 consecutive days. Draining lymph nodes were taken off at 24 hours after the last application. Concentrations: positive control DNCB (dinitrochlorobenzene): 0.5% (w/v) and test item: 10%, 1%, 0.1% (w/v) in DAE 433. Endpoints: ear weight, auricular (ear-draining) lymph node weights and cell counts = lymph node (LN) hyperplasia.


The animals exposed to the test substance at the highest concentration showed skin reactions (congestion of vessels) throughout the experiment. There were no clinical observations attributable to the treatment with test substance at the middle and lower dose level. There was no difference in body weight increment of all groups in comparison to the vehicle control. The positive control substance DNCB elicited a reaction pattern with statistically significant increase in ear weight and LN hyperplasia, which was in congruence with his expected mode of action as a contact allergen. The test substance showed a tendency to increase of ear weight in the highest dose level but without lymph node hyperplasia. Residues of the test substance on the ears caused this increased weight. Comparison of values between treated groups and control group revealed that the test substance did not cause statistically significant increase in LN cell count or in LN weight. Also index of LN weight and LN cell count was not exceeded in any dose level.


In conclusion, at the given experimental conditions the test substance elicited negative result in LLNA test. Based on these results the test substance does not have to be classified as the substance, which may cause sensitisation by skin contact.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
READ ACROSS ANALOGUE APPROACH

The read-across hypothesis is fundamentally based on the same core structure of five ‘yellow disazo condensation pigments’ which optionally can serve as target or as source substances. None of the pigments are sufficiently soluble, either in water or in octanol for systemic uptake or metabolism. The molecular weight ranges from 716.6 g/mol (Pigment Yellow 155) to 1229.2 g/mol (Pigment Yellow 128). Therefore, the molecular weights of all ‘yellow disazo condensation pigments’ are well above the threshold of 500 g/mol, which is generally considered for low dermal and oral uptake [ECHA Guidance R. 7c, 2017]. Furthermore, for each of the substances, the critical body burden (CBB) is above the octanol solubility, which generally indicates a low uptake in biota and makes toxicity unlikely [ECHA Guidance R. 11, 2017].


Please find the complete Read-across justification text including a data matrix and structures attached in the attachment.

Reason / purpose for cross-reference:
read-across source
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
10%
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
none
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
10%
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
none
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
10%
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
10%
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
yes (incl. QA statement)
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
When this study was performed, the LLNA did not yet exist as an OECD testing guideline.
Specific details on test material used for the study:
- yellow powder
- Lot/batch No.: 201P69495
- Expiration date of the lot/batch: 22-MAY-2005
- Stability under test conditions: stable
- Storage condition of test material: room temperature
Species:
guinea pig
Strain:
Himalayan
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: RCC Ltd, Biotechnology & Animal Breeding Division, Wölferstrasse 4, CH-4414 Fullinsdorf
- Age at study initiation: 4 - 6 weeks at delivery
- Weight at study initiation: 300 - 359 g
- Housing: Individually
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: one week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 (air conditioned room)
- Humidity (%): 30 - 70
- Air changes (per hr): 10 - 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2001-07-03 To: 2001-08-03
Route:
intradermal and epicutaneous
Vehicle:
polyethylene glycol
Concentration / amount:
10% (intradermal)
30% (epicutaneous)
Route:
epicutaneous, occlusive
Vehicle:
polyethylene glycol
Concentration / amount:
10% (intradermal)
30% (epicutaneous)
No. of animals per dose:
10 test group
5 control group
Details on study design:
The intradermal induction of sensitization in the test group was performed in the nuchal region with a 10 % dilution of the test item in PEG 300 and in an emulsion of Freund's Complete Adjuvant (FCA) / physiological saline. The epidermal induction of sensitization was conducted for 48 hours under occlusion with the test item at 30% in PEG 300 one week after the intradermal induction. The animals of the control group were intradermally induced
with PEG 300 and FCA/physiological saline and epidermally induced with PEG 300 under occlusion.
Two weeks after epidermal induction the control and test animals were challenged by epidermal application of the test item at 10% in PEG 300 and PEG 300 alone under occlusive dressing.
Challenge controls:
CONTROL GROUP
TONER YELLOW 3GP, 10% in PEG 300 (left flank)
PEG 300 only (right flank)

TEST GROUP
TONER YELLOW 3GP, 10% in PEG 300 (left flank)
PEG 300 only (right flank)
Positive control substance(s):
yes
Remarks:
2-MERCAPTOBENZOTHIAZOLE
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
10%
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
none
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
10%
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
none
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
10%
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
10%
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none

Challenge with vehicle only caused no skin reactions in the control group and test group, respectively.

CONTROL GROUP

No skin reactions were observed in the animals when treated with either PEG 300 only or when treated with the test item at 10% in PEG 300. Yellow discoloration produced by the test item was noted directly after removal of the patch.

TEST GROUP

No skin reactions were observed in the surviving animals when treated with either PEG 300 only or when treated with the test item at 10% in PEG 300. Yellow discoloration produced by the test item was noted directly after removal of the patch. To remove the discoloration all animals were depilated 3 hours prior to challenge reading.

The body weight of the animals was within the range commonly recorded for animals of this strain and age. No clinical signs were observed. To remove the discoloration all animals were depilated 3 hours prior to challenge reading.

One animal died during the study. No cause of death could be identified and no skin reactions were observed. The death is considered to be incidental.

Interpretation of results:
GHS criteria not met
Conclusions:
No toxic symptoms were evident in the guinea pigs of the control or test group. None of the control and test animals showed skin reactions after the challenge treatment at 10 % (w/w) in PEG 300.
Executive summary:

In order to assess the cutaneous allergenic potential, the Maximization-Test was performed in 15 (10 test and 5 control) female albino guinea pigs, in accordance with OECD Guideline No. 406. The intradermal induction of sensitization in the test group was performed in the nuchal region with a 10% dilution of the test item in PEG 300 and in an emulsion of Freund's Complete Adjuvant (FCA) I physiological saline. The epidermal induction of sensitization was conducted for 48 hours under occlusion with the test item at 30% in PEG 300 one week after the intradermal induction. The animals of the control group were intradermally induced with PEG 300 and FCA/physiological saline and epidermally induced with PEG 300 under occlusion. Two weeks after epidermal induction the control and test animals were challenged by epidermal application of the test item at 10% in PEG 300 and PEG 300 alone under occlusive dressing. Cutaneous reactions were evaluated at 24 and 48 hours after removal of the dressing.


No toxic symptoms were evident in the guinea pigs of the control or test group. None of the control and test animals showed skin reactions after the challenge treatment at 10 % (w/w) in PEG 300.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Potential for skin sensitisation has been investigated for two 'yellow disazo condensation pigments'. A guinea pig maximisation assay according to OECD TG 406 has been performed with CAS 68516-73-4 (Clariant, 2001). The maximization test was performed in 15 (10 test and 5 control) female albino guinea pigs. The intradermal induction of sensitisation in the test group was performed in the nuchal region with a 10% dilution of the test item in PEG 300 and in an emulsion of Freund's Complete Adjuvant (FCA) I physiological saline. The epidermal induction of sensitisation was conducted for 48 hours under occlusion with the test item at 30% in PEG 300 one week after the intradermal induction. The animals of the control group were intradermally induced with PEG 300 and FCA/physiological saline and epidermally induced with PEG 300 under occlusion. Two weeks after epidermal induction the control and test animals were challenged by epidermal application of the test item at 10% in PEG 300 and PEG 300 alone under occlusive dressing. Cutaneous reactions were evaluated at 24 and 48 hours after removal of the dressing. No toxic symptoms were evident in the guinea pigs of the control or test group. None of the control and test animals showed skin reactions after the challenge treatment at 10% (w/w) in PEG 300.

This result was confirmed by a local lymph node assay done with CAS 5580-57-4 according to OECD guideline 429 (Synthesia, 2008). In this study the contact allergenic potential of the test item was evaluated with non radioactive measuring of cell proliferation after topical application to female BALB/c mice. Six mice per group were exposed by test and control substances on the dorsum of both ears once a day during 3 consecutive days. Draining lymph nodes were taken off at 24 hours after the last application. Concentrations: positive control DNCB (dinitrochlorobenzene): 0.5% (w/v) and test item: 10%, 1%, 0.1% (w/v) in DAE 433. Endpoints: ear weight, auricular (ear-draining) lymph node weights and cell counts = lymph node (LN) hyperplasia. The animals exposed to the test substance at the highest concentration showed skin reactions (congestion of vessels) throughout the experiment. There were no clinical observations attributable to the treatment with test substance at the middle and lower dose level. There was no difference in body weight increment of all groups in comparison to the vehicle control. The positive control substance DNCB elicited a reaction pattern with statistically significant increase in ear weight and LN hyperplasia, which was in congruence with his expected mode of action as a contact allergen. The test substance showed a tendency to increase of ear weight in the highest dose level but without lymph node hyperplasia. Residues of the test substance on the ears caused this increased weight. Comparison of values between treated groups and control group revealed that the test substance did not cause statistically significant increase in LN cell count or in LN weight. Also index of LN weight and LN cell count was not exceeded in any dose level. In conclusion, at the given experimental conditions the test substance elicited negative result in LLNA test.

Therefore, it is concluded that all 'yellow disazo condensation pigments' are not sensitising to skin and have not to be classified for skin sensitisation.   

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008
The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on available data on skin sensitisation, the test item is not classified according to Regulation (EC) No 1272/2008 (CLP), as amended for the tenth time in Regulation (EU) No 2017/776.