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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
Study period:
Test start date: 15 January, 1990; Test end date: 01 March, 1990; Study completion date: 31 May, 1990
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1990
Report date:
1990

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
adopted May 12, 1981
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
GLP compliance:
no
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Data from a reliable in vivo test conducted before the enforcement of Commission Regulation (EU) 640/2012 of 06 July 2012 amending, for the purpose of its adaptation to technical progress, Regulation (EC) No 440/2008 laying down test methods pursuant to Regulation (EC) No 1907/2006 of the European Parliament and of the Council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) are available.

Test material

Constituent 1
Reference substance name:
Reaction mass of 6-chloro-2-[4-[ethyl(2-hydroxyethyl)amino]phenyl]-1-methylbenz[cd]indolium acetate and 6-bromo-2-[4-[ethyl(2-hydroxyethyl)amino]phenyl]-1-methylbenz[cd]indolium acetate and 6-chloro-2-[4-[ethyl(2-hydroxyethyl)amino]phenyl]-1-methylbenz[cd]indolium hydroxide
IUPAC Name:
Reaction mass of 6-chloro-2-[4-[ethyl(2-hydroxyethyl)amino]phenyl]-1-methylbenz[cd]indolium acetate and 6-bromo-2-[4-[ethyl(2-hydroxyethyl)amino]phenyl]-1-methylbenz[cd]indolium acetate and 6-chloro-2-[4-[ethyl(2-hydroxyethyl)amino]phenyl]-1-methylbenz[cd]indolium hydroxide
Test material form:
solid
Specific details on test material used for the study:
Identified as: FAT 31064/F
Description: Powder
Batch Number: EN 158496.82 / HEW 133/6
Stability of test article: stable; Expiration date: November 1994
Stability of test article dilution: Stable for at least 2 hours
Safety precautions: Gloves, goggles and face mask were sufficient to assure personnel health and safety.

In vivo test system

Test animals

Species:
guinea pig
Strain:
other: Ibm: GOHI; SPF-quality guinea pigs
Sex:
male/female
Details on test animals and environmental conditions:
Test System
The albino guinea pig is the recommended species for skin sennsitization studies.
- Source: BRL, Biological Research Laboratories Ltd., Wolferstrasse 4, CH-4414 Füllinsdorf
- Total Number of Animals per Test: 15 males and 15 females
- Age at acclimatization start: males: 7 weeks, females: 8 weeks
- Body Weight at Acclimatization Start: males: 401 - 479 g; females: 353 - 449 g
- Acclimatization: One week under test conditions after veterinary examination.

Husbandry
- Standard Laboratory Conditions
Air-conditioned with 10-15 air changes per hour and hourly monitored environment with temperature 22 ± 3 degrees centigrade, relative humidity 40-70 %, 12 hours artificial fluorescent light/12 hours dark, music/light period.

- Accommodation: Individually in Makrolon type-3 cages with standard softwood bedding ("Lignocel", Schill AG, CH-4132 Muttenz).

- Diet: Pelleted standard Kliba 342, Batches 55/89 and 56/90 guinea pig breeding/maintenance diet ("Kliba", Klingentalmühle AG, CH-4303 Kaiseraugst), ad libitum.

- Water: Community tap water from Itingen, ad libitum.

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
other: ethanol
Concentration / amount:
5%
Day(s)/duration:
At the start
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
epicutaneous, occlusive
Vehicle:
other: Ethanol
Concentration / amount:
25%
Day(s)/duration:
one week after intradermal induction
Adequacy of induction:
non-irritant substance, but skin pre-treated with 10% SDS
Challengeopen allclose all
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
other: ethanol
Concentration / amount:
25 %
Day(s)/duration:
Two weeks after the epidermal induction
Adequacy of challenge:
highest non-irritant concentration
No.:
#2
Route:
epicutaneous, occlusive
Vehicle:
other: ethanol
Concentration / amount:
25%
Day(s)/duration:
Two weeks after the first challenge
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
10 males and 10 females for test group
5 males and 5 females for control group
Details on study design:
RANGE FINDING TESTS:
A preliminary study was performed to identify irritant test article concentrations suitable for the induction phase of the main study. In addition, a suitable non-irritant concentration of the test article, by the topical route of administration, was identified for the challenge application.

MAIN STUDY
A. INDUCTION EXPOSURE (Intradermal induction)
- No. of exposures: 1
- Test groups: 1
- Control group: 1
- Site: shaved neck
- Frequency of applications: once
- Concentrations: 5%

B. INDUCTION EXPOSURE (Epidermal induction)
- No. of exposures: 1
- Test groups: 1
- Control group: 1
- Site: neck of the animals
- Frequency of applications: once
- Duration of exposure: 48 h
- Concentrations: 25% (with prior application of SDS 10%)

C. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: two weeks after epidermal induction
- Exposure period: 24 hours
- Test groups: 1
- Control group: 1
- Site: flank
- Concentrations: 25%
- Evaluation (hr after challenge): 24 h after removing the dressing

D. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: two weeks after first challenge
- Exposure period: 24 hours
- Test groups: 1
- Control group: 1
- Site: flank
- Concentrations: 25%
- Evaluation (hr after challenge): 24 h after removing the dressing
Positive control substance(s):
yes
Remarks:
Formaldehyde

Results and discussion

Positive control results:
9/10 rats showed positive reactions after first challenge exposure.

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
Intradermal induction: 5%, Epidermal induction: 25%, challenge: 25%
No. with + reactions:
6
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
Intradermal induction: 5%, Epidermal induction: 25%, challenge: 25%
No. with + reactions:
5
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
test chemical
Dose level:
Intradermal induction: 5%, Epidermal induction: 25%, challenge: 25%, rechallenge: 25%
No. with + reactions:
5
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
test chemical
Dose level:
Intradermal induction: 5%, Epidermal induction: 25%, challenge: 25%, rechallenge: 25%
No. with + reactions:
4
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
no indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
no indication of skin sensitisation
Reading:
rechallenge
Hours after challenge:
24
Group:
negative control
Dose level:
vehicle only
No. with + reactions:
0
Total no. in group:
10
Reading:
rechallenge
Hours after challenge:
48
Group:
negative control
Dose level:
vehicle only
No. with + reactions:
0
Total no. in group:
10
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
20% Epidermal, 15% Challenge
No. with + reactions:
9
Total no. in group:
10
Remarks on result:
positive indication of skin sensitisation

Any other information on results incl. tables

Mortality: No death occurred during the study.

Systemic symptoms: No systemic symptoms were observed during the study.

Body weights: The body weight gain of the animals was not affected adversely during the study.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
FAT 31064/F was considered to be not a sensitiser to skin.
Executive summary:

The skin sensitisation potential of FAT 31064/F has been investigated in a study conducted using Magnusson and Kligman maximation method as described in OECD Guideline 406. In this study, a group of guinea pigs containing 10 males and 10 females, received FAT 31064/F at 15 % concentration in the intradermal induction phase and at 25 % concentration during the epidermal induction phase. The day before epidermal induction, the application sites were pretreated with 10 % sodium lauryl sulfate as open application. Two weeks after the epidermal induction application the animals were challenged with the occluded epidermal application for 24 hours on the flank with FAT 31064/F (concentration 25 %) in ethanol and the vehicle alone. A second challenge was performed two weeks after the first challenge. 06/20 and 5/20 animals showed positive reactions at 24-hour and 48-hour readings, respectively, when treated with 25 % test article dilution. No positive reactions were evident with ethanol alone. In the second challenge, 5/20 and 4/20 animals showed positive reactions at 24-hour and 48-hour readings respectively when treated with 25x test article dilutions. No positive reactions with ethanol alone. The positive skin reactions observed in the test group were below 30 % threshold required for the substance to be assessed skin sensitiser. Hence, based on the findings of this study, FAT 31064/F was considered to be not a skin sensitiser.