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Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Remarks:
Buehler assay
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2009-11-17 to 2010-03-23
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report date:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
Type of study:
Buehler test
Justification for non-LLNA method:
The Buehler method is a scientifically well-established OECD guideline method and the results revealed by the Bühler test eventually provide a high level of safety relevant to the workers under realistic conditions (LLNA in some cases tends to give false positive results).
Second, the registrant has always been using the Bühler method for skin sensitization testing for similar substances. Therefore, in this particular case the registrant chose to apply the Bühler test for reasons of comparability of the test results with historical testing data obtained for other substances. Finally, as the test item is registered as an intermediate only, there was no regulatory requirement for testing.

Test material

Constituent 1
Chemical structure
Reference substance name:
2-{[(4R,5R)-2,2,4-trifluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}ethan-1-ol; 2-{[(4R,5S)-2,2,4-trifluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}ethan-1-ol; 2-{[(4S,5R)-2,2,4-trifluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}ethan-1-ol; 2-{[(4S,5S)-2,2,4-trifluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}ethan-1-ol
EC Number:
700-535-6
Cas Number:
1190931-34-0
Molecular formula:
C6F6H6O5
IUPAC Name:
2-{[(4R,5R)-2,2,4-trifluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}ethan-1-ol; 2-{[(4R,5S)-2,2,4-trifluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}ethan-1-ol; 2-{[(4S,5R)-2,2,4-trifluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}ethan-1-ol; 2-{[(4S,5S)-2,2,4-trifluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}ethan-1-ol
Details on test material:
- Name of test material (as cited in study report): DIOX alcohol
- Substance type: organic mono constituent
- Physical state: Colourless liquid
- Analytical purity: no data
- Lot/batch No.: 107-31
- Expiration date of the lot/batch: no data
- Stability under test conditions: no data
- Storage condition of test material: Ambient temperature

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Italia S.p.A., Calco (Lecco), Italy
- Age at study initiation: 4 to 5 weeks old
- Weight at study initiation: 284 to 321 grams
- Housing: Stainless steel cages measuring, with grid floor.
- Diet (e.g. ad libitum): 8GP17, ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C ± 2°C
- Humidity (%): 55% ± 15%
- Air changes (per hr): 15 - 25
- Photoperiod (hrs dark / hrs light): Artificial (fluorescent tubes), daily light/dark cycle of 12/12 hours

IN-LIFE DATES: From: 2009-12-16 To: 2010-01-20

Study design: in vivo (non-LLNA)

Induction
Route:
epicutaneous, semiocclusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% test item concentration for induction and challenge
Day(s)/duration:
6-hr exposure, once a week for 3 weeks
Adequacy of induction:
highest technically applicable concentration used
Challenge
No.:
#1
Route:
epicutaneous, semiocclusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% test item concentration for induction and challenge
Day(s)/duration:
exposure: 6 hours; 1st reading 24h after removal of the dressing; 2nd reading 24h later
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
Test group: 20
Control group: 10
Details on study design:
RANGE FINDING TESTS:
Five animals were used in the preliminary assay to determine the suitable test item concentration; each received 2 concentrations on either flank. The test item was applied at 100, 50, 20, 10 and 5% v/v in sterile water.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 3
- Exposure period: 6 hours
- Test groups: On the day of dosing (Day 1) the hair was clipped from the inductuion site of each animal. Animals of the test group were treated with the undiluted test item. A gauze patch measuring 20x20 mm was soaked with 0.4 ml of the test item and placed onto the selected skin site. All animals of the test group were treated with the test item in this manner and animals of the control group were similarly treated with the selected vehicle (sterile water).
- Control group: see above
- Site: anterior region of the left flank
- Frequency of applications: once weekly
- Duration: 3 weeks
- Concentrations: 100% test item concentration

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 29
- Exposure period: 6 hours
- Test groups: On Day 29, the hair was removed with electric clippers from both the challenge exposure sites of all animals of both test and control groups. A 0.4 ml aliquot of the undiluted test item was spread evenly over an absorbent patch measuring approximately 20x20 mm. This was placed onto the skin of the posterior region of the prepared site on the right flank. A similar patch, containing 0.4 ml of the vehicle selected for the challenge (sterile water), was placed onto the anterior region of the prepared site. All animals of the test and control groups were treated with both the test
item and vehicle in this manner. After an exposure period of approximately 6 hours the dressings were removed and the treated sites cleaned of the remaining test item by washing with lukewarm water.
- Control group: see above
- Site: anterior and posterior regions of the right flank
- Concentrations: 100% test item concentration
- Evaluation (hr after challenge): Approximately 24 hours after removal of the dressing, the treated sites were examined for any signs of reaction to treatment. Skin reaction on the treated sites was again assessed approximately 24 hours after the first examination (approximately 48 hours after removal of the patches).
Challenge controls:
The undiluted test item was selected for the challenge as no irritation was noted in all main phase animals.
Positive control substance(s):
yes
Remarks:
α-Hexylcinnamaldehyde in April 2009 was acceptable; 2-Mercaptobenzothiazole in December 2009/January 2010 was not acceptable (0% response); 2-Mercaptobenzothiazole in February 2010 was acceptable (65% response).

Results and discussion

Positive control results:
Periodical System Reliability check:
A positive control check using alpha-Hexylcinnamaldehyde was performed on April 2009 and the incidence at challenge was acceptable.
A positive control check using 2-Mercaptobenzothiazole (protocol deviation) was performed on December 2009 to January 2010 and the incidence at challenge (0%) was not acceptable. A second positive control check was performed using 2-Mercaptobenzothiazole on February 2010. The incidence at this third positive control was acceptable (65%).

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
100%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
100%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation
Remarks:
Negative control valid
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
0
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation
Remarks:
Negative control valid
Reading:
other: Periodical test system response check
Group:
positive control
Dose level:
Induction: 70% alpha-hexylcinnamaldehyde (HCA) in DMSO; challenge: 15% HCA in acetone
Remarks on result:
positive indication of skin sensitisation
Remarks:
20% response in test group; 0% in control group. Test system valid.
Reading:
other: Periodical test system response check
Group:
positive control
Dose level:
Induction: 50% 2-mercaptobenzothiazole in DMSO; challenge: 50% 2-mercaptobenzothiazole in corn oil
Remarks on result:
no indication of skin sensitisation
Remarks:
0% response in both test and copntrol groups. Test system regarded as invalid. The assay was repeated.
Reading:
other: Periodical test system response check
Group:
positive control
Dose level:
Induction: 50% 2-mercaptobenzothiazole in DMSO; challenge: 50% 2-mercaptobenzothiazole in corn oil
Remarks on result:
positive indication of skin sensitisation
Remarks:
65% response in test group; 0% response in control group at challenge. Test system is valid.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
These results indicate that the test item does not elicit a sensitisation response in the guinea pig, being there no evidence of response at challenge following a period of induction exposure to the test item, DIOX alcohol.
Executive summary:

The potential of the test item, DIOX alcohol, to induce and elicit delayed dermal sensitisation was assessed by a guinea pig model using the methods of Buehler. The concentrations of the test item used in the main study were determined on the basis of the results obtained from a preliminary screening test carried out on DIOX alcohol (maximum concentration tested: 100%). The main sensitisation test was undertaken using a test group of 20 animals and a control group of 10 animals. In an attempt to induce sensitisation, test animals were treated by topical application of the undiluted test item. This was repeated at weekly intervals for a total of 3 weeks. Animals of the control group were treated in the same manner but the vehicle alone (sterile water) was used in place of the test item. Two weeks after the third and final induction exposure, animals of the test and control groups were challenged by topical application of both the vehicle alone (sterile water) and the test item at the same concentration used for induction. At challenge no response was observed to the test item, at the selected concentration, in either test or control group animals. No reaction was observed to the vehicle alone (sterile water). These results indicate that the test item does not elicit a sensitisation response in the guinea pig, being there no evidence of response at challenge following a period of induction exposure to the test item, DIOX alcohol.