Copper, [29H,31H-phthalocyaninato(2-)-N29,N30,N31,N32]-, sulfo [[4-[[2-(sulfooxy)ethyl]sulfonyl]phenyl]amino]sulfonyl derivs.

Endpoint summary

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1989

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant with international guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Species:

guinea pig

Strain:

Pirbright-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: HOECHST AG, Kastengrund, SPF-Zucht
- Strain: Hoe DHPK (SPFLac)
- Weight at study initiation: mean: 291 g (= 100%); mean min: 267 g (-8.2%); mean max: 315 g (+8.2%); n=15
- Housing: in fully air-conditioned rooms in Makrolon cages (type 4) on softwood granules, in groups of 5 animals
- Diet: ERKA-mixed feed No. 8300 for guinea pigs and rabbits, ad libitum
- Water: tap water ad libitum in plastic bottles
- Acclimation period: min. 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 3°C
- Humidity: 50 ± 20 %
- Photoperiod: 12 hours cycle dark/light

Route:

intradermal

Vehicle:

physiological saline

Concentration / amount:

treatment group
25% in 0.9% NaCl solution

Route:

epicutaneous, occlusive

Vehicle:

physiological saline

Concentration / amount:

treatment group
25% in 0.9% NaCl solution

No. of animals per dose:

Determination of the primary concentration: 6
Determining compatibility of intradermal dose: 3
Support Group: 5
Control group: 5
Treatment group: 10

Details on study design:

SUBSTANCE FORMULATIONS
The Freund's Adjuvant was mixed immediately before use with an equal volume of 0.9 % of NaCl solution. This 50 % Freund's Adjuvant preparation was injected intradermal to the animals.

For the dermal application Reactive Blue 21 was diluted with 0.9% (w/v) NaCl.

For the intradermal injections Reactive Blue 21 was dissolved in Freund's adjuvant and diluted with 0.9% NaCl solution pyrogen-free (percentages in w/v) and this solution then diluted with an equal volume of original - mixed Freund's adjuvant .
In the maximization test concentrations cannot be standardized. The suitable concentrations would be determined in preliminary studies. The concentration used depends on the individual test phase.

DETERMINATION OF THE PRIMARY NON-IRRITANT CONCENTRATION
Each of the following test concentrations were administered (occlusive) to the left flank of two guinea pigs:
25 % in 0.9% of NaCl solution
2.5 % in 0.9% of NaCl solution
0.25 % in 0.9% of NaCl solution
The right and left flank of the animals was depilated mechanically. 0.5 mL of the test substance preparation was administered to a 2 x 2 cm cellulose patch, which was fixed to the left flank and covered occlusively for 24 hours with a bandage foil. 24 hours after removal of the patches, the treated skin areas were examined for erythema and edema.

RANGE FINDING TESTS:
To determine the tolerance of intradermal injections, each of the following preparations was administered twice by intradermal injection to 3 guinea pigs. The injection sites (sites 1, 2 and 3) were all within a dorsal area measuring 2 x 4 cm in the vicinity of the shoulders.
site appl. vol. conc. vehicle
1 2x0.1 mL 5.0 % Reactive Blue 21 in 0.9% NaCl solution
2 2x0.1 mL 1.0 % Reactive Blue 21 in 0.9% NaCl solution
3 2x0.1 mL 0.2 % Reactive Blue 21 in 0.9% NaCl solution

MAIN STUDY
Guinea pigs were shaved meccanically over a dorsal area of 4 x 6 cm in the vicinity of the shoulders.

INTRADERMAL INDUCTION EXPOSUR from day 1
- Site: dorsal area of 2 x 4 cm
- Applications: Two intradermal injections per animal and 3 injection sites in a area of 2 x 4 cm. The injection sites were left uncovered.
Groups concentration
site appl. vol. conc. vehicle
1 2x0.1 mL - 50% Freund’s adjuvant solution
2 2x0.1 mL 0.5% Reactive Blue 21 in 0.9% NaCl solution
3 2x0.1 mL 0.5 % Reactive Blue 21 in Freund’s adjuvant solution
Control group
site appl. vol. vehicle
1 2x0.1 mL 50 % Freund’s adjuvant solution
2 2x0.1 mL 0.9% NaCl solution
3 2x0.1 mL 50 % Freund’s adjuvant solution


DERMAL INDUCTION EXPOSUR from day 9
- Concentration: 0.5 mL of the 2.5% test substance preparation
- Site: 2 x 4 cm
- Applications: By cellulose patch. This patch covered the area were intradermal injection had been placed. The administration area was then kept under an occlusive bandage covered with any impermeable film and an elastic bandage for 48 h
- Treatment group: 2.5 % Reactive Blue 21 in 0.9% in NaCl solution
- Control and satellite group: 0.9% NaCl solution
Bandage was removed at day 11

CHALLENGE EXPOSURE from day 22
An area of approx. 5 x 5 cm on the left and right flank was shaved mechanically.
- Concentration: 0.5 mL of the 2.5% test substance preparation
- Site: 2 x 2 cm
- Bandage: the administration area was kept for 24 under an occlusive bandage covered with an impermeable film and an elastic bandage.

At day 23 the occlusive bandage was removed.

Challenge controls:

left flank: 2.5% Reactive Blue 21 in 0.9% NaCl solution

Positive control substance(s):

yes

Remarks:

bi-annulal testing

Reading:

1st reading

Hours after challenge:

48

Group:

test chemical

Dose level:

2.5% test item inside 0.9% NaCl solution

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 48.0. Group: test group. Dose level: 2.5% test item inside 0.9% NaCl solution. No with. + reactions: 0.0. Total no. in groups: 10.0.

Reading:

2nd reading

Hours after challenge:

72

Group:

test chemical

Dose level:

2.5% test item inside 0.9% NaCl solution

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 72.0. Group: test group. Dose level: 2.5% test item inside 0.9% NaCl solution. No with. + reactions: 0.0. Total no. in groups: 10.0.

DETRMINATION OF THE PRIMARY NON-IRRITANT CONCENTRATION

After treatment with 25% substance, one of five animals showed very mild erythema.

The treatment with Freund's Complete Adjuvant may lower the threshold of primary irritation, determined in the preliminary test. Therefore, 5 animals of the satellite group, treated with Freund's Complete Adjuvant, received a dermal administration of 2.5% of Reactive Blue 21.

 

These animals showed no signs of irritation. Therefore, a concentration of 2.5% of Reactive Blue 21 in NaCl solution was chosen for dermal challenge treatment at day 22.

 

DETRMINATION OF THE TOLERANCE OF THE INTRADERMAL INJECTIONS

The intradermal injections with the 5% preparation caused swelling, scaling and induration, the 1% formulation caused a perceivable swelling. Redness did not occur. The injection of the 0.2% formulation resulted no irritation.

Based on this preliminary test, a 0.5% preparation was selected for the intradermal injections in the main test.

 

MAIN TEST

-Body weight gains and clinical signs: The treated animals showed no clinical signs of intoxication throughout the study.

The intradermal injections with Freund's Adjuvant without test substance caused strong redness and swelling at the injection sites, which in case of the Freund's Adjuvant with test substance could not be assessed due to blue discoloration.

Additionally, beginning on day 3 after administration the injection sites were scaled and indurated.

After the removal of the patches on day 11, the injection sites of the animals from the control and satellite group were strongly swollen and necrotic,

the test group additionally discoloured blue.

The body weight gains of the animals were not impaired.

 

-Dermal challenge treatment: 24 hours after removal of the occlusive bandageboth the animals in the control as well as the animals in the treatment group showed no irritation symptoms.

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The present study has shown that after the challenge no animal has given positive response.
According to the evaluation criteria of Directive 83/467/EEC, Reactive Blue 21 is non-sensitizing in the maximization test.

Executive summary:

Sensitisation was assessed using the Magnusson and Kligman sensitisation assay. No irritation reactions were noted in both control and test groups. Under the conditions of the present study, none of ten animals of the treatment group showed a positive skin response after the challenge procedure.  

On the basis of the results, the substance cannot be considered to be a skin sensitiser.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Sensitisation was assessed using the Magnusson and Kligman sensitisation assay. No irritation reactions were noted in both control and test groups. Under the conditions of the present study, none of ten animals of the treatment group showed a positive skin response after the challenge procedure.  

On the basis of the results, the substance is considered not to be a skin sensitiser.

Migrated from Short description of key information:
No skin sensitizing properties observed

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available