Registration Dossier

Toxicological information

Skin sensitisation

Currently viewing:

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2002-04-11 to 2002-05-10
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 guideline
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
1996
Deviations:
yes
Remarks:
unclear how long the epidermal induction period lasted;
GLP compliance:
yes (incl. QA statement)
Remarks:
signed 2001-08-22
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
According to the REACH regluation (EC) No. 1907/2006, the LLNA test is the first-choice method for in vivo testing and in exceptional circumstances another test can be used. Since the study was carried out before the regulation entered into force and the guinea pig maximisation test according to OECD 406 is an acceptable method for testing skin sensitisation, it is not justified to conducted an additional LLNA test due to animal welfare.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Details on test material:
- State of aggregation: yellow green powder
Specific details on test material used for the study:
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: dry

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelmann GmbH, Gartenstraße 27, D-33178 Borchen, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: < 1 year
- Weight at study initiation: 255 - 326 g
- Housing: housed individually Macrolon cages (Surface: 780 cm²)
- Diet: Raiffeisen Ringfutter, Trockenfutter 52 P Raiffeisen, Kraftfutter Kehl
- Water (ad libitum): tap water (additive: 20 mg ascorbic acid and 100 mg citric acid per 100 mL water)
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 20 - 23 °C
- Relative humidity: 30 - 70 %
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Concentration / amount:
0.1 mL of Freund's complete adjuvant (diluted 1:1 with aqua ad iniectabilia)
Day(s)/duration:
Day 0
Adequacy of induction:
non-irritant substance, but skin pre-treated with 10% SDS
Route:
intradermal
Vehicle:
other: sesame oil
Concentration / amount:
0.1 mL of 5 % concentration
Day(s)/duration:
Day 0
Adequacy of induction:
non-irritant substance, but skin pre-treated with 10% SDS
Route:
intradermal
Vehicle:
other: sesame oil
Concentration / amount:
0.1 mL of 5 % test item in a 1:1 mixture (v/v) Freund's complete adjuvant/aqua ad iniectabilia
Day(s)/duration:
Day 0
Adequacy of induction:
non-irritant substance, but skin pre-treated with 10% SDS
Route:
epicutaneous, occlusive
Vehicle:
other: sesame oil
Concentration / amount:
50 % test item
Day(s)/duration:
Day 7
Challenge
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
other: sesame oil
Concentration / amount:
50 % test item
Day(s)/duration:
Day 21 (exposure duration: 24 hours)
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
10 (test substance) + 5 (controls)
Details on study design:
RANGE FINDING TESTS:
Six Dunkin-Hartley guinea pigs were used for the preliminary test: 3 animals for the topical administration and 3 animals for the intracutaneous administration.

a) Intracutaneous administration:
The back of the animals was shaved. Two intradermale injections (100 µL each) of the same test item concentrations were injected on each side of the spinal column. The test material was mixed with sesame oil. Three concentrations were tested during the preliminary test (1, 3, and 5 % of the test item). The injection side were investiged for erythema and oedema 24, 48, and 72 hours after administration.

Results:
No erythema or oedema was observed at any concentration. Therefore, the 5 % concentration was used in the main study even no skin irritation was observed.

b) Topical administration:
Three test item concentrations were used for the topical administration (20, 33, and 50 % (m/v) of the test item). The test material was mixed with sesame oil. The back of the animals was shaved and the test substances was applied onto a patch that was placed on the back. The patch was fixed with occlusive dressing for a duration of 24 hours. After the 24 hour exposure period, the skin was examined for erythema and oedema.

Results:
No erythema or oedema was observed at any concentration. Therefore, the 50 % (m/v) concentration was chosen for testing in the main study.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 (intradermal injection and dermal application)
- Site: back (surface: 4 x 6 cm²)
- Frequency of applications: three pairs of intradermal injections were given in the back region once and 6 days later the skin was treated with sodium lauryl sulfate 10 % in vaseline in order to induce a local irritatrion. 24 hours later the test item was topically applied using a patch-test technique. The patch (2 x 4 cm²) was soaked in the test item (saturated solution of the test item in a few drops of sesame oil), which was placed onto the shaved back of the animals. The patch was covered with a occlusive dressing.
- Concentrations:
Test animals:
Intradermal:
i) 0.1 mL FCA diluted 1:1 with aqua ad iniectabilia
ii) 0.1 mL test item in sesame oil (final concentration: 5 %)
iii) 0.1 mL test item in FCA mixed with aqua ad iniectabilia in the ratio 1:1 (final concentration: 5 %)
Topical application:
50 % concetration of test item

Control animals:
Intradermal injection:
i) 0.1 mL FCA diluted 1:1 with aqua ad iniectabilia
ii) 0.1 mL sesame oil
iii) 0.1 mL sesame oil and FCA/aqua ad iniectabilia (1:1) in the ratio 1:1
Topical application:
sesame oil

B. CHALLENGE EXPOSURE
- No. of exposures: 1 (two weeks after the topical application (induction))
- Exposure period: 24 hours
- Site (test animals and controls): both flanks shaved; right flank: sesame oil; left flank: 50 % of the test item in sesame oil
- Evaluation (hr after challenge): 24 and 48 hours

OTHER OBSERVATIONS:
- mortality, clinical signs, and body weight (at the start of testing and at the end of testing)
Challenge controls:
5 guinea pigs were used as control animals.
Challenge dose: right flank: sesame oil; left flank: 50 % of the test item in sesame oil
Positive control substance(s):
yes
Remarks:
Benzocaine (study conducted from 2002-04-12 to 2002-05-11; 10 test animals and 5 control animals; intradermal (induction): 5 %; topical (induction): 100 %; challenge: 100 %)

Results and discussion

Positive control results:
Animals treated with benzocaine in sesame oil exhibited a sensitising reaction in 6 of 10 animals in form of a discrete/patchy or moderate and confluent erythema (grades 1 and 2) 24 hours after administration. Furthermore, 4 of 10 animals exhibited discrete or patchy erythema (grade 1) 48 hours after administration.

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
50 % test item
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No mortality, clinical signs or skin reactions were observed. No effect on body weight was observed.
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
50 % test item
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No mortality, clinical signs or skin reactions were observed. No effect on body weight was observed.
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
50 % test item
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
No mortality, clinical signs, or skin reactions were observed. No effect on body weight was observed.
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
50 % test item
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
No mortality, clinical signs, or skin reactions were observed. No effect on body weight was observed.
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
100 % concentration
No. with + reactions:
6
Total no. in group:
10
Clinical observations:
No mortality of clinical signs were observed. No effect on body weight was observed.
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
100 % concentration
No. with + reactions:
4
Total no. in group:
10
Clinical observations:
No mortality of clinical signs were observed. No effect on body weight was observed.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
According to the Regulation (EC) 1272/2008 and subsequent adaptations, the substance is not sensitising to the skin.