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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:
19 January 2005 to 31 March 2005
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Guideline-conform study under GLP without deviations.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005
Report date:
2005

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
1992
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
1996
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of study:
Buehler test
Justification for non-LLNA method:
The Buehler test was conducted prior to the change in Regulation and data requirements.

Test material

Constituent 1
Chemical structure
Reference substance name:
2-[difluoro(methoxy)methyl]-1,1,1,3,3,3-hexafluoropropane
EC Number:
609-534-4
Cas Number:
382-26-3
Molecular formula:
C5H4F8O
IUPAC Name:
2-[difluoro(methoxy)methyl]-1,1,1,3,3,3-hexafluoropropane
Constituent 2
Reference substance name:
Propane, 1, 1, 1, 3, 3- pentafluoro- 3- methoxy- 2- (trifluoromethyl) –
IUPAC Name:
Propane, 1, 1, 1, 3, 3- pentafluoro- 3- methoxy- 2- (trifluoromethyl) –
Test material form:
other: liquid
Details on test material:
- Name of test material (as cited in study report): Octafluoroisobutyl methyl ether
- Substance type: pure substance
- Physical state: liquid
- Analytical purity: >86%
- Impurities (identity and concentrations): not reported
- Composition of test material, percentage of components: not reported
- Isomers composition: not reported
- Purity test date: not reported
- Lot/batch No.: 20/02/04-161
- Expiration date of the lot/batch: 2006
- Stability under test conditions: not reported
- Storage condition of test material: in the refrigerator (range of 5 +- 3 °C)

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male

Study design: in vivo (non-LLNA)

Induction
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
INDUCTION: 100% (undiluted)
Day(s)/duration:
1 patch/week for 3 weeks
Adequacy of induction:
highest technically applicable concentration used
Challenge
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
polyethylene glycol
Concentration / amount:
CHALLENGE: 5% in Polyethylene glycol
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
20 test animals + 10 control animals
Details on study design:
RANGE FINDING TESTS:
Irritation Screen I: 4 concentrations were tested: 100%, and 75%, 50%, 25% (w/w) diluted in PEG300
Irritation screen II: 4 concentrations were tested: 15%, 10%, 5% and 3% (w/w) in PEG300

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 1 patch/week over 3 weeks (3 exposures)
- Exposure period: 1 application per week, for 6 hrs
- Test groups: 20 males
- Control group: 10 males
- Site: 25 mm Hill Top chamber applied on the left shoulder of each test animal
- Frequency of applications: once a week
- Duration: 3 weeks
- Concentrations: undiluted substance

B. CHALLENGE EXPOSURE
- No. of exposures: one application
- Day(s) of challenge: 2
- Exposure period: 6 hrs
- Test groups: 20 males
- Control group: 10 males
- Site: left posterior quadrant of the side and back of the animals
- Concentrations: 5% in PEG 300
- Evaluation (hr after challenge): 24 and 48 hrs
Challenge controls:
Ten animals of the control group were not treated during the induction and were challenged with the test substance at 5% in PEG 300.
Positive control substance(s):
yes
Remarks:
5% alpha-Hexylcinnamaldehyde, in a separate run within 6 month prior

Results and discussion

Positive control results:
1st challenge: 3% negative
2nd challenge: 5% => 7/20 positive responses at 24h and 11/20 positive responses at 48h

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
5% in PEG 300
No. with + reactions:
7
Total no. in group:
19
Clinical observations:
One animal was found dead on test day 25. No clinical signs were observed on the other animals
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
5% in PEG 300
No. with + reactions:
6
Total no. in group:
19
Clinical observations:
No clinical effects observed
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
5% alpha-Hexylcinnamaldehyde in PEG 300
No. with + reactions:
7
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Remarks:
Re-challenge at a slightly higher concentration
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
5% alpha-Hexylcinnamaldehyde in PEG 300
No. with + reactions:
11
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Remarks:
Re-challenge at a slightly higher concentration

Any other information on results incl. tables

ERYTHEMA SCORE TEST GROUPS CONTROL GROUP
20 animals 10 animals
24 hrs 48 hrs 24 hrs 48 hrs
1 12 13 10 10
2 7 6 0 0
3 0 0 0 0
4 0 0 0

0

No. with grades >= 1 0 0 0 0
No. tested  19 19 10 10
INCIDENCE* 7/19 *** 0/10
SEVERITY** 0.3 - 0.4 0
* number of animals showing a response of grade 1 or greater at either 24- or 48- hour reading out of the total animals.
** Total sum of 24- and 48-hours response readings divided by the number of animals exposed (maximum of 3).
*** One animal of the test group was found dead on day 25 (i.e. 10 days after the last induction). At necropsy, congestion in the lung was noted. The death may be related to the test item treatment, however, the single incidence in one of the 20 tested animals, may also be regarded as spontaneous and treatment unrelated.

Grading of animals was done by positioning each animal under true light.

The grading method used for irritation screens, induction and challenge was identical.

The scoring system was performed by visual assessment of erythema, oedema and other clinical changed in skin conditions. They were assessed as following:

0 no visible change
1 discrete or patchy erythema
2 moderate or confluent erythema
3 intense erythema and swelling

Applicant's summary and conclusion

Interpretation of results:
Category 1B (indication of skin sensitising potential) based on GHS criteria
Conclusions:
2-[difluoro(methoxy)methyl]-1,1,1,3,3,3-hexafluoropropane meets the Regulation EC No. 1272/08 classification criteria for Skin Sensitisation Sub-Category 1B.
Executive summary:

The purpose of the study was to assess the possible allergenic potential of 2-[difluoro(methoxy)methyl]-1,1,1,3,3,3-hexafluoropropane when administered topically to Albino Dunkin Hartley guinea pigs. For this purpose the "Buehler test" was used.

Twenty male animals of the test group were treated topically with 2-[difluoro(methoxy)methyl]-1,1,1,3,3,3-hexafluoropropane at 100% once a week for a 3 -week induction phase. Two weeks after the final induction application the animals were challenged with the test item concentration of 5% in Polyethylene glycol (PEG) 300. The ten animals of the control group were not treated during the induction. They were treated once at challenge with 2-[difluoro(methoxy)methyl]-1,1,1,3,3,3-hexafluoropropane at 5% in PEG 300.

After challenge treatment, 37% (at the 24 -hour reading) and 32% (at the 48 -hours reading) of the animals of the test group were observed with skin reactions. No skin reactions were observed in the control group treated in the same conditions during the challenge phase.

According to Regulation (EU) No. 1272/08, a substance has to be classified as skin sensitizer if a response of at least 15 % of the animals in a non-adjuvant guinea pig test method is observed. Furthermore, in accordance to the second adaptation to technical progress for the CLP Regulation, if in the Buehler test there is a positive response between 15 % and 60 % responding at > 0,2 % to ≤ 20 % topical induction dose or a positive response ≥ 15 % at > 20 % topical induction dose the substance has to be classified as Skin Sensitizer Sub-Category 1B.

2-[difluoro(methoxy)methyl]-1,1,1,3,3,3-hexafluoropropane meets the above reported classification criteria and therefore it needs to be classified as skin sensitizer Sub-Category 1B.

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