Sodium 6,14-diethyl-1,1,1,2,2,18,18,19,19,19-decafluoro-8,12-dioxo-10-({[1-(2,2,3,3,3-pentafluoropropoxy)butan-2-yl]oxy}carbonyl)-4,7,13,16-tetraoxanonadecane-9-sulfonate

Endpoint summary

Description of key information

Key, skin sensitisation, in vivo, mice, OECD 429, GLP: no sensitiser

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 APR - 2 AUG 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Version / remarks:

22 JUL 2010

Deviations:

yes

Remarks:

During the acclimation phase the relative humidity was between 36 - 65 % instead of 45 - 65%. Furhtermore, the different test item concentrations were prepared individually instead of serially.

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Version / remarks:

30 MAY 2008

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

CBA

Remarks:

CBA/CaOlaHsd

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories B.V. Postbus 6174 NL-5960 AD Horst
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 10 – 11 weeks
- Weight at study initiation: 21.6 ± 1.4 g
- Housing: Grouped in Makrolon Type II cages with wire mesh top and granulated soft wood bedding
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: ≥ 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 45 – 65%
- Air changes (per hr): About 10 / hour
- Photoperiod (hrs dark / hrs light): 12 hrs dark / 12 hrs light
- IN-LIFE DATES: From: day 1 To: day 5

Vehicle:

dimethylformamide

Concentration:

5, 10 and 25 %

No. of animals per dose:

5

Details on study design:

COMPOUND SOLUBILITY
The highest test item concentration, which can be technically used, was a 50 % (w/w) solution in dimethylformamide.

PRE-SCREEN TESTS:
To determine the highest non-irritant test concentration that did at the same time not induce signs of systemic toxicity, a pre-test was performed. Two animals were treated by application of test item concentrations of 25 and 50% (w/w) once daily each on three consecutive days. At the tested concentrations the animals did not show any signs of systemic toxicity. From day 2 to 6, the treated animals showed an erythema of ear skin (animal treated with 25%: day 2 to 6 erythema Score 1, animal treated with 50%: day 2, day 5, and day 6 erythema Score 2, and day 3 as well as day 4 erythema Score 3). Additionally, hair loss was observed from day 3 to day 6. Due to the signs of excessive local skin irritation noted at 50% (w/w) test item concentration, 25% (w/w) was chosen as highest test item concentration.

MAIN STUDY

ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response:
A test item is regarded as a sensitiser in the LLNA if the following criteria are fulfilled:
- First, that exposure to at least one concentration of the test item resulted in an incorporation of ³H-methyl thymidine (³HTdR) at least 3-fold or greater than that recorded in control mice, as indicated by the Stimulation Index.
- Second, that the data are compatible with a conventional dose response, although allowance must be made (especially at high topical concentrations) for either local toxicity or immunological suppression.

TREATMENT PREPARATION AND ADMINISTRATION:
Each test group of mice was treated by topical (epidermal) application to the dorsal surface of each ear with different test item concentrations of 5, 10, and 25% (w/w) in dimethylformamide. The application volume, 25 µL/ear/day, was spread over the entire dorsal surface (average ~ 8 mm) of each ear once daily for three consecutive days. A further group of mice was treated with an equivalent volume of the relevant vehicle alone (control animals).

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Statistics:

- The mean values and standard deviations were calculated in the body weight tables and for the DPM values (group mean DPM ± standard deviation).
- The Dean-Dixon-Test and Grubb’s test were used for identification of possible outliers (performed with Microsoft Excel 2003). One outlier was identified by means of the Dean Dixon test only.
- The ANOVA (Dunnett-test) was conducted on the DPM values and ear weights to assess whether the difference was statistically significant between test item groups and negative control (vehicle) group. However, both biological and statistical significance were considered together.

Positive control results:

Conc. Si
0% 1.00
5% 0.88
10% 1.51

Parameter:

SI

Value:

1

Test group / Remarks:

0% vehicle control group

Parameter:

SI

Value:

1.16

Test group / Remarks:

5%

Parameter:

SI

Value:

0.77

Test group / Remarks:

10%

Parameter:

SI

Value:

1.5

Test group / Remarks:

25%

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
0% vehicle control group: 714.2 mean DPM/animal
5% group: 829.4 mean DPM/animal
10% group: 552.8 mean DPM/animal
25% group: 1068.2 mean DPM/animal

DETAILS ON STIMULATION INDEX CALCULATION
Mean DPM/animal was determined by dividing the sum of the measured values from lymph nodes of all animals (2 lymph nodes/animal) within a group by the number of animals in that group (5 animals)

EC3 CALCULATION
The EC3 value could not be calculated, because all SIs are below the threshold value of 3 for a positive response.

CLINICAL OBSERVATIONS:
No signs of systemic toxicity were observed during the study period. From day 2 to 5, the animals showed an erythema of the ear skin (animals treated with 5%: on day 3 and day 4 erythema Score 1, animals treated with 10%: from day 2 to 4 erythema Score 1, animals treated with 25%: day 2 (within 1 hour prior to the 2nd application), day 4 and day 5 erythema Score 1, and additionally, on day 2 (within 1 hour after the 2nd application) and day 3 erythema Score 2).

BODY WEIGHTS
The body weight of the animals, recorded prior to the first application and prior to treatment with ³HTdR, was within the range commonly recorded for animals of this strain and age.

Viability / mortality

No deaths occurred during the study period.

Ear weight

The measured ear weight of all animals treated was recorded on day 6 after necropsy. A statistically significant or biologically relevant increase in ear weight was not observed in any test item treated group in comparison to the vehicle control group.

Interpretation of results:

other: EU GHS criteria not met

Conclusions:

It can be concluded that the test item was not a skin sensitiser under the test conditions of this study.

Executive summary:

Study design

This GLP study was performed according to OECD Guidelines for Testing of Chemicals, No. 429: Skin Sensitisation: Local Lymph Node Assay (adopted on 22 July 2010). In the study the test item dissolved in dimethylformamide was assessed for its possible skin sensitising potential in mice. For this purpose a local lymph node assay was performed using test item concentrations of 5, 10, and 25% (w/w). The highest concentration tested was the highest concentration that could be applied whilst avoiding local skin irritation as confirmed by a pre-experiment.

 

Results

The animals did not show any signs of systemic toxicity during the course of the study and no cases of mortality were observed. From day 2 to 5, the animals showed an erythema of the ear skin (animals treated with 5%: on day 3 and day 4 erythema Score 1, animals treated with 10%: from day 2 to 4 erythema Score 1, animals treated with 25%: day 2 (within 1 hour prior to the 2^nd application), day 4 and day 5 erythema Score 1, and additionally, on day 2 (within 1 hour after the 2^nd application) and day 3 erythema Score 2). A statistically significant or biologically relevant increase in ear weight was not observed in any test item treated group in comparison to the vehicle control group. Stimulation Indices (S.I.) of 1.16, 0.77, and 1.50 were determined with the test item at concentrations of 5, 10, and 25% (w/w) in dimethylformamide, respectively. A statistically significant or biologically relevant increase in DPM values was not observed in any test item treated group in comparison to the vehicle control group and a dose response was also not observed. An outlier was identified in the vehicle control group (DPM value determined for animal number 4). However, as exclusion of the outlier did not change the overall test result, the value in question was not excluded from calculation.

 

Conclusion

The test item was thus not a skin sensitiser under the test conditions of this study.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Skin sensitisation

Skin sensitisation, OECD 429

The skin sensitisation potential of the test item was determined in a test following GLP and according to OECD Guidelines for Testing of Chemicals, No. 429: Skin Sensitisation: Local Lymph Node Assay (adopted on 22 July 2010).

Study design

In the study the test item dissolved in dimethylformamide was assessed for its possible skin sensitising potential in mice. For this purpose a local lymph node assay was performed using test item concentrations of 5, 10, and 25% (w/w). The highest concentration tested was the highest concentration that could be applied whilst avoiding local skin irritation as confirmed by a pre-experiment.

 

Results

The animals did not show any signs of systemic toxicity during the course of the study and no cases of mortality were observed. From day 2 to 5, the animals showed an erythema of the ear skin (animals treated with 5%: on day 3 and day 4 erythema Score 1, animal streated with 10%: from day 2 to 4 erythema Score 1, animals treated with 25%: day 2 (within 1 hour prior to the 2^nd application), day 4 and day 5 erythema Score 1, and additionally, on day 2 (within 1 hour after the 2^nd application) and day 3 erythema Score 2). A statistically significant or biologically relevant increase in ear weight was not observed in any test item treated group in comparison to the vehicle control group. Stimulation Indices (S.I.) of 1.16, 0.77, and 1.50 were determined with the test item at concentrations of 5, 10, and 25% (w/w) in dimethylformamide, respectively. A statistically significant or biologically relevant increase in DPM values was not observed in any test item treated group in comparison to the vehicle control group and a dose response was also not observed. An outlier was identified in the vehicle control group (DPM value determined for animal number 4). However, as exclusion of the outlier did not change the overall test result, the value in question was not excluded from calculation.

 

Conclusion

The test item was thus not a skin sensitiser under the test conditions of this study.

Justification for classification or non-classification

Based on the results of the key study for in vivo skin sensitisation, the registered substance is not subject to classification as skin sensitiser in accordance with Regulation (EC) No 1272/2008.