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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
adopted 1992
Deviations:
no
GLP compliance:
yes
Type of study:
Buehler test
Justification for non-LLNA method:
The current accepted and preferred method for skin sensitisation testing according to the REACH legislation (EC no 1907/2006) and CLP Regulation (EC No 1272/2008) is the murine local lymph node assay (LLNA). A validated test method, OECD TG 429 (OECD 2002) is available for the LLNA. The guideline acknowledges the limits of the LLNA, and states that there are instances where test substance classes or substances containing functional groups shown to act as potential confounders make the use of guinea pig tests more appropriate. It is concluded that the LLNA is not applicable where the properties of the test material cause interference in the accuracy of the LLNA (OECD 2002). The statement in the OECD TG 429 is given with reference to the findings of Basketter et al. (2009a), who demonstrated false positives in silicone based substances which had previously been demonstrated to be non-sensitisers in the guinea pig maximisation test (GPMT). The importance of available evidence from guinea pig results, consideration of chemical reactivity, epidermal bioavailability and clinical and experimental human data are emphasised as central to reaching appropriate regulatory decisions for substances which have been shown to fall outside the specificity of the LLNA (Basketter et al., 2009b). The non-applicability of the LLNA for silicone based substances has also been demonstrated by Petry et al. (2012). The sensitisation potential of polyfunctional silicone materials was tested in a comparative study investigating the GPMT and the LLNA assays, which found the five tested substances to be negative in the GPMT whereas they were concluded to be weak to moderate skin sensitisers in the LLNA (Petry et al., 2012).

Test material

Constituent 1
Chemical structure
Reference substance name:
-
EC Number:
449-160-7
EC Name:
-
Cas Number:
116912-64-2
Molecular formula:
C7H18N2O4Si C8H20N2O4Si C9H22N2O4Si C10H24N2O4Si
IUPAC Name:
[3-(ethoxydimethoxysilyl)propyl]urea; [3-(trimethoxysilyl)propyl]urea; {3-[diethoxy(methoxy)silyl]propyl}urea
Details on test material:
- Name of test material (as cited in study report): A-1160 (methanol stripped)
- Physical state: clear, colourless liquid
- Substance type: Alkoxysilane
- Storage condition of test material: At room temperature in the dark under nitrogen

In vivo test system

Test animals

Species:
guinea pig
Strain:
Himalayan
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Biotechnology and animal breeding division (RCC Ltd.), Fuellinsdorf, Switzerland
- Age at study initiation: approx. 4 weeks
- Weight at study initiation: 294 ± 14 g (control group), 308 ± 16 g (experimental group)
- Housing: 5 animals per metal cage
- Diet: Standard guinea pig diet including ascorbic acid (1000 mg/kg); Charles River breeding and maintenance diet for guinea pigs, Altromin, Lage, Germany, ad libitum
- Water: Tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 3
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod: 12 hrs dark / 12 hrs light

Study design: in vivo (non-LLNA)

Induction
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% (0.5 ml of undiluted test substance (experimental group); 0.5 ml of water (control group))
Challenge
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% (0.15 ml of undiluted test substance (experimental group); 0.15 ml of water (control group))
No. of animals per dose:
Control group: 10 animals
Experimental group: 20 animals
Details on study design:
RANGE FINDING TESTS:
In a range finding test the application of undiluted test substance to 2 animals did not reveal any erythema or edema findings 24 and 48 h after exposure.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 3 (day 1, 8, and 15)
- Exposure period: 6 h
- Test groups: 0.5 ml undiluted test substance
- Control group: 0.5 ml water
- Site: left side of the scapular region
- Frequency of applications: every 7 days
- Duration: day 1-15
- Concentrations: 100%

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day of challenge: 28
- Exposure period: 6 h
- Test groups: 0.15 ml undiluted test substance + 0.15 mL water
- Control group: 0.15 ml undiluted test substance + 0.15 mL water
- Site: right flank
- Concentrations: 50%
- Evaluation (hr after challenge): 24, 48
Challenge controls:
The control group is actually a challenge control.
Positive control substance(s):
no
Remarks:
The sensitivity of the test system was checked at least every 6 months.

Results and discussion

Positive control results:
No positive control was used with this study, but data are available showing that a reliability check is carried out at regular intervals. The positive control substance (Alpha-hexylcinnamic aldehyde) was used for a reliability check, and induced positive reactions in 60% of the animals, thus meeting the reliability criteria for the Buehler test (≥ 15% positive response).

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
50%
No. with + reactions:
0
Total no. in group:
10
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
50%
No. with + reactions:
0
Total no. in group:
20
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
50%
No. with + reactions:
0
Total no. in group:
10
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
50%
No. with + reactions:
0
Total no. in group:
20
Group:
positive control
Remarks on result:
positive indication of skin sensitisation

Applicant's summary and conclusion

Interpretation of results:
other: CLP/EU GHS criteria are not met, no classification required according to Regulation (EC) No 1272/2008
Conclusions:
In a Buehler Test conducted to GLP and OECD 406 (reliability score 1) ureidopropyltrialkoxysilane, mixed methoxy and ethoxy esters was not sensitising to the skin.