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Administrative data

Description of key information

Skin sensitisation (OECD 406, guinea pig): not sensitising

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:
25 Jul - 07 Sep 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
17 July 1992
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Test was done before LLNA as first-choice method for in-vivo testing was set into force.
Species:
guinea pig
Strain:
other: Crl: HA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River, Sulzfeld, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 5 weeks
- Weight at study initiation: 355 - 420 g
- Housing: animals were kept in groups in Terluran-cages on Altromin saw fibre bedding (lot no. 300312).
- Diet: autoclaved hey and Altromin 3122 maintenance diet for guinea pigs (rich in crude fibre, lot no. 1725), ad libitum
- Water: tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 10
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12
Route:
intradermal
Vehicle:
physiological saline
Concentration / amount:
1.5%
Day(s)/duration:
single injection
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
epicutaneous, occlusive
Vehicle:
other: Vaseline
Concentration / amount:
50%
Day(s)/duration:
2
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
other: Vaseline
Concentration / amount:
3%
Day(s)/duration:
1
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
5 (controls), 10 (test groups)
Details on study design:
RANGE FINDING TESTS: a preliminary test with 7 animals was performed to determine appropriate treatment concentrations for the main assay. Cutaneous reactions (erythema and edema) were evaluated 24, 48 and 72 h after injection or topical application with the test substance.

By intradermal route:
- 1 animal: intradermal administrations of the test substance emulsified in physiological saline at concentrations of 2.5 and 5%
- 1 animal: intradermal administrations of the test substance emulsified in physiological saline at concentrations of 1 and 1.5%

By cutaneous route;
- 1 animal: topical treatment with the test substance dissolved in vaseline at concentrations of 50 and 100% for 24 h
- 1 animal: topical treatment with the test substance dissolved in vaseline at concentrations of 50 and 100% for 48 h
- 1 animal: topical treatment with the test substance dissolved in vaseline at concentrations of 12.5 and 25% for 24 h
- 1 animal: topical treatment with the test substance dissolved in vaseline at concentrations of 12.5 and 6% for 48 h
- 1 animal: topical treatment with the test substance dissolved in vaseline at concentrations of 3 and 6% for 24 h

Scored erythema and edema are listed in Table 1 under “Any information on materials and methods incl. tables”.

Based on the results (mild-to-moderate skin irritation, but well tolerated systemically), the following concentrations were chosen for the main assay:
- intradermal induction: 1.5%; epicutaneous induction: 6%; challenge application: 3%

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 (intradermal and epicutaneous, respectively)
- Exposure period: single injection (intradermal) and 48 h (epicutaneous)

- Test groups:
Intradermal (3 pairs of injections):
Injection 1: a 1:1 mixture (v/v) FCA/physiological saline 0.9% NaCl
Injection 2: test substance in physiological saline 0.9% NaCl
Injection 3: test substance in a 1:1 mixture (v/v) FCA/ physiological saline 0.9% NaCl
Epicutaneous: test substance in vaseline

- Control group:
Intradermal (3 pairs of injections):
Injection 1: a 1:1 mixture (v/v) FCA/physiological saline 0.9% NaCl
Injection 2: 100% vaseline
Injection 3: 50% (v/v) formulation of vaseline in a 1:1 mixture (v/v) FCA/ physiological saline 0.9% NaCl
Epicutaneous: vaseline

- Site: shoulder region (intradermal and epicutaneous)
- Frequency of applications: twice (Day 0 and Day 7)
- Duration: 0-7
- Concentrations: intradermal 1.5%, epicutaneous 6%

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 20
- Exposure period: 24 h
- Test groups: test substance and vaseline only
- Control group: test substance and vaseline only
- Site: left flank (test substance) and right flank (vehicle = intraspecific control)
- Concentrations: 3%
- Evaluation (hr after challenge):24 and 48 h after patch removal

OTHER:
On Day 6 (approx. 24 h before epicutaneous application of the test substance), the test area of the animals was clipped and they were treated with 0.5 g of 10% sodium lauryl sulphate in vaseline in order to induce local irritation.
Challenge controls:
The control group is actually a challenge control.
Positive control substance(s):
yes
Remarks:
mercaptobenzothiazole (2 and 15% (w/w))
Positive control results:
The sensitivity and reliability of the experimental technique was routinely checked using the positive control substance mercaptobenzothiazole. Ten animals were intradermally (2%) and epicutaneously (25%) induced with the positive control substance, followed by challenge treatment at 15% concentration. The sensitisation rate after challenge application of the positive control substance was 100%, thus fulfilling the criteria for a positive result (≥ 30%) and verifying the sensitivity and reliability of the assay.
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
induction: 0%; challenge: 3%
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
induction: 1.5 and 6%; challenge: 3%
No. with + reactions:
1
Total no. in group:
10
Clinical observations:
1/10: erythema grade 1
Remarks on result:
other: slight evidence of sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
induction: 2 and 25%; challenge: 15%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
3/10: erythema grade 1; 5/10: erythema grade 2; 2/10: erythema grade 3; 1/10: edema grade 1; 1/10: edema grade 2
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
induction: 0%; challenge: 3%
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
induction: 1.5 and 6%; challenge: 3%
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
induction: 2 and 25%; challenge: 15%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
2/10: erythema grade 1; 5/10: erythema 2; 3/10: erythema grade 3; 1/10: edema grade 1; 1/10: edema grade 2
Remarks on result:
positive indication of skin sensitisation

For the test item, at the first reading time point following challenge, i.e., 24 h, 1/10 animals showed skin reaction described as skin erythema graded 1. In the study report, the finding was described as slight sensitisation. At the second reading time point, i.e., 48 h, none of the 10 animals showed any skin reaction. Thus, under the test conditions used, no obvious skin sensitizing potential could be evidenced with respect to the test item. The suitability of the test system as well as the reliability of the method both were confirmed by the results of the positive control.

Interpretation of results:
other: CLP/EU GHS criteria not met, no classification required according to Regulation (EC) No. 1272/2008
Conclusions:
CLP: not classified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

There is a reliable guinea pig maximisation test (GPMT) available for Amines C12-14-branched alkyl, monohexyl and dihexyl phosphates (CAS 96690-34-5) in order to fulfill the standard data requirements defined in Regulation (EC) No 1907/2006, Annex VIII, 8.3.

The Guinea pig maximisation test was performed with the test substance according to the OECD guideline 406 and under GLP conditions (BSL Bioservice, 2012). In the main study, 10 female guinea pigs were treated with the test substance at 1.5% in physiological saline for intradermal induction on day 1 and at 6% in vaseline for epicutaneous induction on day 7. Five animals served as negative controls. 20 days after the epidermal induction, epidermal challenge was performed with 3%. Mercaptobenzothiazole was used as positive control substance in the study and was tested within a routine check 6 months before the main test. 10 of 10 animals of the positive control group showed skin reactions 24 and 48 h after the challenge. With respect to the test group, skin examination after 24 h following challenge revealed slight skin erythema in one of the 10 animals. After 48 h, none of the 10 animals showed any skin reaction. Thus, the test material was found to be not sensitising to the skin of guinea pigs under the test conditions used.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The available data on the skin sensitising potential of the substance do not meet the criteria for classification according to Regulation (EC) 1272/2008, and are therefore conclusive but not sufficient for classification.