reaction mass of: tetrasodium 7-(4-(4-fluoro-6-(4-(2-sulfonatoethylsulfonyl)phenylamino)-1,3,5-triazin-2-ylamino)-2-ureidophenylazo)naphthalene-1,3,6-trisulfonate;tetrasodium 7-(4-(4-hydroxy-6-(4-(2-sulfonatoethylsulfonyl)phenylamino)-1,3,5-triazin-2-ylamino)-2-ureidophenylazo)naphthalene-1,3,6-trisulfonate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27 January 1998 to 27 February 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

study was already available

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

427-650-1

EC Name:

-

IUPAC Name:

reaction mass of: tetrasodium 7-(4-(4-fluoro-6-(4-(2-sulfonatoethylsulfonyl)phenylamino)-1,3,5-triazin-2-ylamino)-2-ureidophenylazo)naphthalene-1,3,6-trisulfonate and tetrasodium 7-(4-(4-hydroxy-6-(4-(2-sulfonatoethylsulfonyl)phenylamino)-1,3,5-triazin-2-ylamino)-2-ureidophenylazo)naphthalene-1,3,6-trisulfonate

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): Reaktivgelb FD 08064

In vivo test system

Test animals

Species:

guinea pig

Strain:

Pirbright-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: HARLAN WINKELMANN
- Weight at study initiation: mean - 366g (320 - 399g)
- Housing: in fully air-conditioned rooms in macrolon cages (type 4) on soft wood granulate in groups of 5 animals
- Diet (e.g. ad libitum): ssniff® Ms-H (V2233), ad libitum
- Water (e.g. ad libitum): tap water in plastic bottles, ad libitum
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 (± 3°C)
- Humidity (%): 50 (± 20 %)
- Photoperiod (hrs dark / hrs light): 12 hours daily

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

water

Remarks:

deionizied

Concentration / amount:

The following preparations were used for the intradermal injections of 0.1 mL each:

Control group
1.) 50 % Freund's Complete Adjuvant emulsion: Original Freund's Complete Adjuvant (Sigma Chemical Company) was mixed immediately before use with an equal volume of deionized water.
2.) Deionized water
3.) 50 % Freund's Complete Adjuvant emuision mixed with an equal volume of the vehicle

Treatment group
1.) 50 % Freund's Complete Adjuvant emulsion: Original Freund's Complete Adjuvant (Sigma Chemical Company) was mixed immediately before use with an equal volume of deionized water.
2.) 5 % Reaktivgelb FD 08064 in deionized water
3.) 5 % Reaktivgelb FD 08064 in a 50 % Freund's Complete Adjuvant emulsion

For the intradermal injections of the test substance in 50 % Freund's adjuvant, Reaktivgelb FD 08064 was dissolved in deionized water and then mixed with an equal volume of Freund's Original Adjuvant [percentages w/v].

Day(s)/duration:

Day 1

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Induction 2

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

25% / 0.5 mL

Day(s)/duration:

Day 8/48 h

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

water

Remarks:

deionizied

Concentration / amount:

5%/0.5 mL

Day(s)/duration:

Day 22/24 h

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

Test group Number of animals
Determination of the primary non-irritant concentration 3
Determination of the tolerance of the intradermal injections 2
Control group 5
Treatment group 10

10 animals in the treatment group and 5 animals in the control group were used. In case of a questionable result additional animals will be tested to give a total of 20 test and 10 control animals.

Details on study design:

RANGE FINDING TESTS:

Prior to the determination of the primary non-irritation concentration in a dermal-occlusive test the animals received 4 intradermal injections of a 50% Freund's Complete Adjuvant emulsion (4 x 0.1 ml) into the dorsal area, since Freund's Complete Adjuvant may lower the threshold of primary irritation. Thereafter, each of the following test concentrations was administered to the flanks of two guinea pigs:

25.0 % in deionized water
5.0 % in deionized water
1.0 % in deionized water

The hair on the flanks of the animals was removed mechanically. 0.5 ml of the test substance preparation was administered to a 2 x 2 cm cellulose patch, which was fixed to the flank and covered occlusively for 24 hours with a bandage and film. 24 hours after removal of the patches, the treated skin areas were examined for erythema and oedema.

Determination of the tolerance of the intradermal injections

To determine the tolerance of intradermal injections, each of the following prepara¬tions was administered twice by intradermal injection to 2 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.
24, 48, 72 and 96 hours after administration the injection sites were examined for local tolerance.


MAIN STUDY

A. INDUCTION EXPOSURE
- No. of exposures: 6
- Exposure period: Intradermal 7 days, dermal 48 hours
- Test groups: 50% Freund's Adjuvant, 5% substance in deionised water, 5% substance with 50% Freund's Adjuvant
- Control group: 50% Freund's Adjuvent, deinioised water, equal volume of deionised water with 50% Freund's Adjuvent
- Site: Within dorsal area
- Frequency of applications: once

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 22
- Exposure period: 24 hours
- Test groups: Test substance
- Control group: Test substance
- Site: Left flank
- Concentrations: 5.0%
- Evaluation (hr after challenge): 24 & 48 hours

Main test

Chronological description of the test procedure indicating the day, at which procedure was carried out, on the left margin of the page.
Intradermal induction treatment

Two intradermal injections per animal of the following preparations. The injection sites (site 1, 2 and 3) were all within a dorsal area of 2 x 4 cm (see prior page). The injection sites were left uncovered.

Treatment group:

site appl. vol. vehicle
1 2x0.1 ml' - 50 % Freund's Adjuvant
2 2x0.1 ml 5% substance in deionized water
3 2x0.1 ml 5 % substance in 50 % Freund's Adjuvant

Control group:
site appl. Vol. vehicle
1 2 x 0.1 ml 50 % Freund's Adjuvant
2 2x0.1 ml deionized water
3 2 x 0.1 ml equal volume of deionized water and 50 % Freund's Adjuvant

The administration area was examined for local tolerance. Any systemic toxic effects were recorded.

Dermal induction treatment

An amount of 0.5 ml of the test substance preparation (treatment group) or the vehicle (control group) was administered to a 2 x 4 cm cellulose patch. This patch covered the area where the intradermal injection had been placed. The administration area was then kept for 48 hours under an occlusive bandage with an impermeable film and an elastic bandage.
Treatment group:25.0 % test substance in deionized water
Control group: deionized water

Dermal challenge treatment
One area of approx. 5 x 5 cm on the left flank was shaved mechanically.

An amount of 0.5 ml of the test substance preparation was administered to a 2 x 2 cm cellulose patch. The administration area was then kept for 24 hours under an occlusive bandage with an impermeable film and an elastic bandage.

Treatment and control group (left flank): 5.0 % Reaktivgelb FD 08064 in deionized water
After treatment, the occlusive bandage removed. Any remnants of the test substance were carefully washed off with warm water. Examination of the skin was undertaken approx. 24 and 48 hours after removal of the patches. Body weights of the test animals determined.

Positive control substance(s):

yes

Results and discussion

Positive control results:

valid

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Remarks on result:

positive indication of skin sensitisation

Results 2

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

5%

No. with + reactions:

0

Total no. in group:

5

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

no indication of skin sensitisation

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

5%

No. with + reactions:

0

Total no. in group:

5

Results 5

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

no indication of skin sensitisation

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the present study, none of ten animals of the treatment group showed a positive skin response after the challenge procedure.

Based on the results of this study Reaktivgelb FD 08064 showed no evidence for sensitizing properties.

Executive summary:

Study conducted to EU test guidance 96/54/EEC part B6 and OECD test guideline 406 in compliance with GLP.

The substance showed no evidence for sensitizing properties.