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Diss Factsheets

Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002
Report date:
2002

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
1992
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
1996
GLP compliance:
yes (incl. QA statement)
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
An in vitro or in chemico skin sensitisation study does not need to be conducted because adequate data from an in vivo skin sensitisation study (different from LLNA test) is available.

Test material

Constituent 1
Chemical structure
Reference substance name:
-
EC Number:
444-290-0
EC Name:
-
Cas Number:
508202-43-5
Molecular formula:
Not applicable
IUPAC Name:
tricobalt(3+) dodecasodium tris(2-[2-(2-oxido-5-sulfamoylphenyl)diazen-1-yl]-6-(phenylamino)-3-sulfonatonaphthalen-1-olate) tris(2-[2-(2-oxido-5-sulfonatophenyl)diazen-1-yl]-6-(phenylamino)-3-sulfonatonaphthalen-1-olate)

In vivo test system

Test animals

Species:
guinea pig
Strain:
Himalayan
Remarks:
Ibm: GOHI
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
-Source: RCC Ltd, Biotechnology & Animal Breeding Division, Wölferstrasse 4. CH-4414 Füllinsdorf / Switzerland
-Age at study initiation: 4 - 6 weeks
-Weight at study initiation:
Pretest groups: 426 - 448 g
Control and test group 372 - 433 g
-Housing: individually in Makrolon type-4 cagas with standard softwood bedding ("Lignocel". Schiil AG, CH-4132 Muttenz).
-Diet: Pelleted standard Provimi Kliba 3418, guinea pig breeding / maintenance diet, containing Vitamin C (Provimi Kiiba AG, CÍH-4303 Kaiseraugst), ad libitum.
-Water: Community tap water from Füilinsdorf, ad libitum.
-Acclimation period: one week (no acclimatization for the animals of the pretest)
-Indication of any skin lesions: only animals without any visible signs of illness were used for the study.

ENVIRONMENTAL CONDITIONS
-Temperature: 20 ± 3 °'C
-Humidity: 30-70 %
-Air changes: 10-15 alr changes per hour
-Photoperiod: 12 hours cycle dark/light
-Other: music was played during the daytime light period

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
propylene glycol
Remarks:
PEG 300
Concentration / amount:
20 %
Day(s)/duration:
a single injection
Route:
epicutaneous, semiocclusive
Vehicle:
propylene glycol
Remarks:
PEG 300
Concentration / amount:
50 %
Day(s)/duration:
48 hours
Challenge
No.:
#1
Route:
epicutaneous, semiocclusive
Vehicle:
polyethylene glycol
Remarks:
PEG 300
Concentration / amount:
15 %
Day(s)/duration:
24 hours
No. of animals per dose:
PRE-TEST
1 male for intradermal induction at concentration of 20 %,15 % and 10 %
2 males for epidermal application at concentration of 50 %, 25 %,15 % and 10 %

MAIN TEST
10 males for intradermal induction at concentration of 20 %, epidermal application of 50 % and challenge dose of 15 %
5 males for control group trated only with vehicle (induction dose) and challenge with vehicle only (right flank) and dose of 15 % (left flank)
Details on study design:
PRETEST
I) INTRADERMAL INJECTIONS:
-No. of exposures: 3 ijections
-Exposure period: before the study initiation date
-Test groups: one guinea pig
-Site: shaved neck
-Concentrations:
A = 20 %, B = 15 % and C = 10 %
-Other: dermal reactions were assessed 24 hours later.
II) EPIDERMAL APPLICATIONS:
-No. of exposures: 4 patches
-Exposure period: before the study initiation date
-Test groups: two guinea pigs.
-Site: shaved flank
-Concentrations:
D = 50 %, E = 25 %, F = 15 % and G = 10 %
-Other: the reaction sites were assessed 24 and 48 hours after removal of the bandage for erythema and oedema

MAIN STUDY
Based on the results obtained the concentration selected for the main study were:
-for intradermal induction 20 %
-for induction and challenge in the main study 50 % and 15%, respectively.

I) INDUCTION EXPOSURE
-No. of exposures: 3 pairs of intradermal injections (0.1 ml/site)
-Test groups: 10 males guinea pig
-Control group: 5 males guinea pig
-Site: scapular region
-Concentrations: the test item at 20 %

II) EPIDERMAL EXPOSURE
-No. of exposures: one application
-Exposure period: 48 hours
-Test groups: 10 males guinea pig
-Control group: 5 males guinea pig
-Site: scapular area
-Concentrations: test item (50 % in PEG 300)
-Evaluation: the reaction sites were assessed 24 and 48 hours after removal of the bandage

CHALLENGE EXPOSURE
-No. of exposures: one application
-Exposure period: 24 hours
-Test groups: 10 males guinea pig
-Control group: 5 males guinea pig
-Site: left and right flank
-Concentrations: 15 % applied to the left flank and the vehicle only was applied on the right flank
-Evaluation: the reaction sites were assessed 24 and 48 hours after removal of the bandage.
Challenge controls:
yes
Positive control substance(s):
yes
Remarks:
Alpha-hexylcinnamaldehyde (tested in a current reference study)

Results and discussion

Positive control results:
No toxic symptoms were evident in the guinea pigs of the control or test group. No deaths occurred. All test animals (at the 24-hour reading) and 6 out of 10 animals (at the 48-hour reading) showed discrete/patchy erythema after the challenge treatment with Alpha-hexylcinnamaldehyde at 0.1 % (w/w) in PEG 300. No skin effect was observed in the control group.

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
15 %
No. with + reactions:
10
Total no. in group:
10
Key result
Reading:
1st reading
Hours after challenge:
48
Group:
test chemical
Dose level:
15 %
No. with + reactions:
8
Total no. in group:
10
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
15 %
No. with + reactions:
0
Total no. in group:
5
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
15 %
No. with + reactions:
0
Total no. in group:
5
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
0.1 %
No. with + reactions:
10
Total no. in group:
10
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
0.1 %
No. with + reactions:
6
Total no. in group:
10

Any other information on results incl. tables

SKIN EFFECTS AFTER INTRADERMAL INDUCTION

The expected and common findings were observed in the control and test group after the different applications using FCA intradermally. These findings consisted of erythema, oedema, necrotizing dermatitis, encrustation, exfoliation of encrustation and also violet discoloration produced by the test item.

No detailed description of the effects is given in the report as these FCA effects are well known.

SKIN EFFECTS AFTER EPIDERMAL INDUCTION

-CONTROL GROUP: no erythematous or oedematous reaction was observed in the animals treated with PEG 300 only.

-TEST GROUP: ss the test item at 50 % stained the skin violet, it was not possible to determine whether erythema was present or not. However, no oedema was observed. The animals were not depilated in the epidermal induction phase.

SKIN EFFECTS AFTER THE CHALLENGE

-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 15 % in PEG 300.

Violet 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.

-TEST GROUP: discrete/patchy to moderate/confluent erythema were observed in all animals at the 24-hour reading and in eight out of 10 animals at the 48-hour reading after treatment with the test Item at 15% in PEG 300. No skin reactions were observed in the animals treated with PEG 300 only. Violet 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.

VIABILITY / MORTALITY / MACROSCOPIC FINDINGS

There were no deaths during the course of the study, hence no necropsies were performed.

CLINICAL SIGNS, SYSTEMIC

No signs of systemic toxicity were observed in the animals.

BODY WEIGHTS

Animals no. 469 of the control group and nos. 481-482 of the test group showed a loss of body weight (0.5 % to 2.6 %) during the acclimatization period. They recovered between the treatment start and the end of the study. The body weight of the other animals was within the range commonly recorded for animals of this strain and age.

Applicant's summary and conclusion

Interpretation of results:
other: Classified according to the CLP Regulation (EC 1272/2008) Skin sens. Cat. 1B
Conclusions:
The substance has sensitising properties.
Executive summary:

In order to assess the cutaneous allergenic potential of test substance, the Maximization-Test was performed in 15 (10 test and 5 control) male albino guinea pigs, in accordance with OECD Guideline No. 406 (1992) and the method B.6 of EEC-Directive 96/54 EEC.

The intradermal induction of sensitization In the test group was performed in the nuchal region with a 20 % 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 50 % 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 15 % in PEG 300 and PEG 300 alone under occlusive dressing. Cutaneous reactions were evaluated at 24 and 48 hours after removal of the dressing.

All test animals (at the 24-hour reading) and eight (at the 48-hour reading) out of 10 test animals showed showed positive reactions after the challenge treatment with test item at 15% (w/w) in PEG 300. No skin effect was observed in the control group.