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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
Study period:
1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
None

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1994
Report Date:
1994

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
Principles of method if other than guideline:
Guidelines followed
GLP compliance:
yes (incl. certificate)
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

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
Identification: FAT 20063/D TE
Lot: BS-BOP 01-15
Appearance: Dark blue powder
Specific details on test material used for the study:
Identification: LANASOL BLAU 8G ROH TROCKEN (FAT 20'063/C)
Description: Dark-blue powder
Batch Number: 3.46
Purity / Formulation: 81.9%
Stability of Test Article: Stable in storage condition; expiration date: 04/99
Stability of Test Article in Vehicle: Unknown in bi-distilled water and in a 1:1 (v/v) mixture of FCA/physiological saline, is excluded from the Statement of Compliance
Storage Conditions: At room temperature (approx. 20°C), away from direct sunlight
Safety precautions: Gloves, goggles and face mask were obligatory to ensure personnel health and safety.

In vivo test system

Test animals

Species:
guinea pig
Strain:
other: Ibm: GOHI; SPF-quality guinea pigs (synonym: Himalayan spotted) delivered by BRL
Sex:
female
Details on test animals and environmental conditions:
Source: BRL, Biological Research Laboratories Ltd. Wölferstrasse 4, 4414 Füllinsdorf/Switzerland
Number of animals for main study / pretest: 30 females/6 females, nulliparous and non-pregnant
Age at beginning of Acclimatization period: 5 - 7 weeks
Body Weight at beginning of Acclimatization period: Control and Test Group 279 - 343 g, Pretest 284 - 354 g
Identification: By unique cage number and corresponding ear tags.
Randomization: Randomly selected at time of delivery.
Acclimatization: One week for the control and test group under test conditions after health examination. No acclimatization for the animals of the pretest. Only animals without any visual signs of illness were used for the study.

Standard Laboratory Conditions:
Air-conditioned with 10-15 air changes per hour and continuously monitored environment with a temperature between 20 - 22 degrees centigrade, a relative humidity between 50 - 80 %, 12 hours artificial fluorescent light (approx. 100 Lux) /12 hours dark, music during the light period.

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

Diet:
Pelleted standard Kliba 342, Batch nos. 64/94 (from 19-SEP-1994) and 65/95 (from 20-SEP-1994) guinea pig breeding/ maintenance diet ("Kliba", Klingentalmühle AG, CH-4303 Kaiseraugst), ad libitum.

Water:
Community tap water from Füllinsdorf, ad libitum. Once weekly additional supply of ascorbic acid (1 g/L) via the drinking water.

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
other:
Concentration / amount:
Freund's complete adjuvant and physiological saline (1:1)
Day(s)/duration:
Test day 1
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
epicutaneous, occlusive
Vehicle:
other: bidistilled water
Concentration / amount:
25 % (highest tested concentration in a pre-test)
Day(s)/duration:
test day 8
Adequacy of induction:
non-irritant substance, but skin pre-treated with 10% SDS
Challenge
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
other: bidistilled water
Concentration / amount:
25 % (highest tested concentration in a pre-test)
Day(s)/duration:
day 22
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
Ten females were used as control group and 20 females were used as test group.
Positive control substance(s):
yes
Remarks:
2-MERCAPTOBENZOTHIAZOL in a separate study "PROJECT 371485" and BENZOCAINE in a separate study " PROJECT 371430"

Results and discussion

Positive control results:
1) In the study with 2-MERCAPTOBENZOTHIAZOL, 60 % of the animals were positive after treatment with a nonirritant test substance concentration of 25 % in mineral oil.
2) Similarly, in the study with BENZOCAINE, 60 % of the animals were positive after treatment with a nonirritant test substance concentration of 25 % in mineral oil

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
25%
No. with + reactions:
0
Total no. in group:
19
Clinical observations:
No toxic symptoms were evident in the guinea pigs of the control or test group.
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
25%
No. with + reactions:
0
Total no. in group:
19
Clinical observations:
No toxic symptoms were evident in the guinea pigs of the control or test group.

Any other information on results incl. tables

RESULTS

Main Study

SKIN EFFECTS AFTER INTRADERMAL INDUCTION PERFORMED ON TEST DAY 1

CONTROL GROUP

Injection site 1: (1:1 (v/v) mixture of Freund's Complete Adjuvant [FCA] and physiological saline)

The area around the injection site was oedematous and erythematous from test day 2 to 4, and then became necrotic from test day 5 to 8 followed by encrustation and exfoliation of encrustation up to the termination of test.

Injection site 2 (bi-distilled water)

The area around the injection site was oedematous and erythematous from test day 2 to 4.

Injection site 3 (1:1 (w/w) mixture of bi-distilled water in a 1:1 (v/v) mixture of FCA and physiological saline)

The reactions observed were identical to those obtained at injection site 1 with the mixture of FCA and physiological saline. As the animals were bandaged with the semi-occlusive dressing no observations of the skin were possible on test day 9.

TEST GROUP

Injection site 1 (1:1 (v/v) mixture of Freund's Complete Adjuvant [FCA] and physiological saline)

The reactions observed were identical to those obtained in the control group with the mixture of FCA and physiological saline, at injection site 1.

Injection site 2 (5 % dilution of test article in bi-distilled water)

The area around the injection site was oedematous from test day 2 to 4 and blue discolored from test day 2 to 8. Necroses were observed from test day 5 to 8 followed by encrustation and exfoliation of encrustation up to the termination of test.

Injection site 3 (5% dilution of test article in a 1:1 (v/v) mixture of FCA and physiological saline)

The reactions observed were identical to those obtained at injection site 2 with the 5 % dilution of test article in bi-distilled water.

As the animals were bandaged with the semi-occlusive dressing no observations of the skin were possible on test day 9.

SKIN EFFECTS AFTER EPIDERMAL INDUCTION PERFORMED ON TEST DAY 8

CONTROL GROUP:

No erythematous or oedematous reaction was observed in the animals treated with bi-distilled water only.

TEST GROUP:

As the test article stained the skin blue, it was not possible to determine whether erythema was present. However, no oedema was observed.

Blue discoloration was noted from test day 10 to 23.

All animals of the control and test group were pretreated with a 10 % SLS in paraffinum perliquidum.

SKIN EFFECTS AFTER THE CHALLENGE PERFORMED ON TEST DAY 22

CONTROL and TEST GROUP:

No positive reactions were observed in the animals either when treated with bidistilled water alone or when treated with the test article at 25 % in bidistilled water. Blue discoloration was noted from test day 23 (after removal of the dressing) to 25.

VIABILITY / MORTALITY / MACROSCOPIC FINDINGS

One animal (no. 343) of the test group was found dead on test day 10. At necropsy a red brown discoloration of the lungs was observed.

CLINICAL SIGNS, SYSTEMIC

No symptoms of systemic toxicity were observed in the animals.

BODY WEIGHTS

One animal of the test group lost weight during the treatment period.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The substance can be considered as not a skin sensitiser.
Executive summary:

The sensitisation potential of the test substance was evaluated in a study conducted according to the maximisation test described in OECD Guideline 406 and EU Method B.6, in compliance with GLP. Ten females were used as control group and 20 females were used as test group. Intradermal and epidermal inductions were carried out on day 1 and day 8 at 5 and 25 % test concentrations, respectively. The test area was pretreated with 10 % Sodium-Lauryl-Sulfate (SLS) in paraffinum perliquidum before epidermal induction. The test and control guinea-pigs were challenged two weeks after the epidermal induction application. The test and control guinea-pigs were treated in the same way. The test concentration used for challenge was 25 %. Approximately 24 and 48 hours after the removal of the dressing the application sites were assessed for erythema and oedema using the numerical scoring system according to Draize. No positive reactions were observed in the animals at 24 and 48 hours after challenge either when treated with bidistilled water alone or when treated with the test article at 25 % in bidistilled water. Hence, the substance can be considered as not a skin sensitiser.