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Description of key information

The test item was considered to be a non-sensitiser under the conditions of the test.

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:
from 2011-12-16 to 2012-03-26
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline-conform study performed under GLP
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
adopted 22 July 2010
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
other: CBA/Ca (CBA/CaOlaHsd)
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories UK Ltd., Oxon, UK
- Age at study initiation: 8 to 12 weeks
- Weight at study initiation: 15 to 23 g
- Housing: individually housed in suspended solid-floor polypropylene cages furnished with softwood woodflakes
- Diet (e.g. ad libitum): 2014C Teklad Global Rodent diet ad libitum
- Water (e.g. ad libitum): mains tap water ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25
- Humidity (%): 30 to 70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12
Vehicle:
dimethylformamide
Concentration:
25 μL of the test item at a concentration of 25%, 10% and 5% w/w in dimethyl formamide per ear (50 µL per animal)
No. of animals per dose:
Vehicle control: 4
5 % w/w: 4
10 % w/w: 4
25 % w/w: 4
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: soluble at 25 % w/w in dimethyl formamide
- Irritation: Yellow coloured residual test item on the ears and fur was noted. No signs of systemic toxicity or visual local skin irritation were noted. A greater than 25% increase in mean ear thickness was noted. As no other signs of irritation were noted the increase was considered to be due to the residual test item on the ears and, although the mean ear thickness increase was greater than 25%, the concentration was selected for the main
test.
- Lymph node proliferation response: not measures

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: LLNA
- Criteria used to consider a positive response: The proliferation response of lymph node cells was expressed as the number of radioactive disintegrations per minute per lymph node (disintegrations per minute/node) and as the ratio of 3HTdR incorporation into lymph node cells of test nodes relative to that recorded for the control nodes (Stimulation Index). The test item will be regarded as a sensitiser if at least one concentration of the test item results in a threefold or greater increase in 3HTdR incorporation compared to control values. Any test item failing to produce a threefold or greater increase in 3HTdR incorporation will be classified as a "non-sensitiser".

TREATMENT PREPARATION AND ADMINISTRATION:
The test item was freshly prepared as a solution in dimethyl formamide. The test item was formulated within two hours of being applied to the test system. It is assumed that the formulation was stable for this duration.
The mice were treated by daily application of 25 μl of the appropriate concentration of the test item to the dorsal surface of each ear for three consecutive days (Days 1, 2, 3). The test item formulation was administered using an automatic micropipette and spread over the dorsal surface of the ear using the tip of the pipette. Five days following the first topical application of the test item or vehicle (Day 6) all mice were injected via the tail vein with 250 μl of phosphate buffered saline (PBS) containing 3H-methyl thymidine (3HTdR: 80 μCi/ml, specific activity 2.0 Ci/mmol, ARC UK Ltd) giving a total of 20 μCi to each mouse.
Positive control substance(s):
other: no positive control
Key result
Parameter:
SI
Remarks:
at 5; 10 and 25%
Value:
>= 1.38 - <= 2.15
Test group / Remarks:
test groups with dose at 5%, 10% and 25% w/w
Remarks on result:
other: 5 % w/w: 1.38 10 % w/w: 1.52 25 % w/w: 2.15
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: Vehicle: 982.42 dpm/Node 5 % w/w: 1360.44 dpm/Node 10 % w/w: 1495.08 dpm/Node 25 % w/w: 2112.12 dpm/Node
Parameter:
SI
Remarks:
10%
Value:
ca. 1.52
Test group / Remarks:
Test group at 10%
Parameter:
SI
Remarks:
25%
Value:
2.15
Test group / Remarks:
Test group at 25%

Clinical Observations and Mortality Data

Yellow coloured residual test item on the ears and fur was noted in animals treated with the test item at a concentration of 25% w/w in dimethyl formamide.

There were no deaths. No signs of systemic toxicity were noted in the test or control animals during the test.

Bodyweight

One animal treated with dimethyl formamide alone showed a greater than expected bodyweight loss. Bodyweight changes of the remaining test animals between Day 1 and Day 6 were comparable to those observed in the corresponding control group animals

over the same period.

Interpretation of results:
GHS criteria not met
Conclusions:
The test item was considered to be a non-sensitiser under the conditions of the test.
Executive summary:

A study was performed to assess the skin sensitisation potential of the test item in the CBA/Ca strain mouse following topical application to the dorsal surface of the ear. The method was designed to be compatible with OECD TG 429.

Following a preliminary screening test in which no clinical signs of toxicity were noted at a concentration of 25% w/w, this concentration was selected as the highest dose investigated in the main test of the Local Lymph Node Assay. Three groups, each of four animals, were treated with 50 μl (25 μl per ear) of the test item as a solution in dimethyl formamide at concentrations of 25%, 10% or 5% w/w. A further group of four animals was treated with dimethyl formamide alone.

The Stimulation Index expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control group are as follows:

Concentration (% w/w) in dimethyl formamide

Stimulation Index

Result

5

1.38

Negative

10

1.52

Negative

25

2.15

Negative

The test item was considered to be a non-sensitiser under the conditions of the test.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

A study was performed to assess the skin sensitisation potential of the test item in the CBA/Ca strain mouse following topical application to the dorsal surface of the ear. The method was designed to be compatible with OECD TG 429.

Following a preliminary screening test in which no clinical signs of toxicity were noted at a concentration of 25% w/w, this concentration was selected as the highest dose investigated in the main test of the Local Lymph Node Assay. Three groups, each of four animals, were treated with 50 μl (25 μl per ear) of the test item as a solution in dimethyl formamide at concentrations of 25%, 10% or 5% w/w. A further group of four animals was treated with dimethyl formamide alone.

The Stimulation Index expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control group are as follows:

Concentration (% w/w) in dimethyl formamide

Stimulation Index

Result

5

1.38

Negative

10

1.52

Negative

25

2.15

Negative

The test item was considered to be a non-sensitiser under the conditions of the test.


Migrated from Short description of key information:
The test item was considered to be a non-sensitiser under the conditions of the test.

Justification for selection of skin sensitisation endpoint:
The selected key study was performed under GLP and in accordance with OECD TG 429. No other studies are available.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available
Additional information:

A study was performed to assess the skin sensitisation potential of the test item in the CBA/Ca strain mouse following topical application to the dorsal surface of the ear. The method was designed to be compatible with OECD TG 429.

Following a preliminary screening test in which no clinical signs of toxicity were noted at a concentration of 25 % w/w, this concentration was selected as the highest dose investigated in the main test of the Local Lymph Node Assay. Three groups, each of four animals, were treated with 50 μl (25 μl per ear) of the test item as a solution in dimethyl formamide at concentrations of 25 %, 10 % or 5 % w/w. A further group of four animals was treated with dimethyl formamide alone.

The Stimulation Index expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control group are as follows:

Concentration (% w/w) in dimethyl formamide

Stimulation Index

Result

5

1.38

Negative

10

1.52

Negative

25

2.15

Negative

The test item was considered to be a non-sensitiser under the conditions of the test.

No information about respiratory sensitisation is available.

Justification for classification or non-classification

The substance does not need to be classified for sensitisation according to Regulation (EC) No 1272/2008 (CLP) because the stimulation index observed in a LLNA was below 3.

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