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Description of key information

Skin Sensitisation:

Two in vivo studies are available.

Local Lymph Node Assay (LLNA): It was conducted using method similar to OECD429. The highest SI (Stimulation Index) of 4.1 was calculated for 25% group and the test item was considered to have an EC3 value of 3%.

An in vivo skin sensitisation study was carried out in accordance with GB/T 21608-2008 which is similar to the BT method of OECD406.

Erythema and/or edema were not be found for the negative control and treatment group at 24 and 48 hours, after challenge and rechallenge exposure, the both sensitization rates were 0%.

Respiratory Sensitisation: No study available.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 2010-03-31 to 2010-04-14
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable study report which meets basic scientific principles. 3 animals per group.
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
adopted in Jul 2010
Deviations:
yes
Remarks:
3 animals per group
GLP compliance:
no
Remarks:
no more details
Type of study:
mouse local lymph node assay (LLNA)
Specific details on test material used for the study:
- Analytical purity: 100%
- Lot/batch No.: 10SC8156928-2-2
Species:
mouse
Strain:
other: CBA/JNCrlj
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Japan Inc.
- Age at study initiation: approximately 9 weeks
- Weight at study initiation: 19.87 - 25.18 g (range)
- Housing: animals were housed 3 per cage in suspended aluminium cages with stainless steel mesh front and floor, w176 x d302 x h130 mm (Yamato Scientific Co., Ltd., Tokyo, Japan). Cages were exchanged with washed, sterile cages every week.
- Diet: CRF-1 diet (Oriental Yeasy Co., Ltd., Japan), ad libitum
- Water: filtered tap water via an automatic water supply equipment (Labo Engineering Inc.), ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-25
- Humidity (%): 40 - 70
- Air changes (per hr): more than 10
- Photoperiod (hrs dark / hrs light): 12/12
Vehicle:
dimethyl sulphoxide
Concentration:
1, 5, 25% (w/v)
No. of animals per dose:
3 females
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: the highest concentration that could be technically used was a 25% suspension in DMSO.
- Irritation: Three mice (1 per dose group) were treated by epidermal application to the dorsal surface of each ear with 1, 5 and 25% concentrations of the test item once daily for 3 consecutive days from Day 0. The ears were assessed for skin irritation on Day 1, 2 and 5. On Day 5, the draining auricular lymph nodes from each ear were excised and weighed. No signs of local irritation was observed in the animals. No treatment-related effects on body weight were noted. The doses applied were selected as the doses to be used in the main study.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: 3H-methyl thymidine incorporation determined by β-scintillation.

- Criteria used to consider a positive response:
The test substance was considered to be positive for skin sensitisation when the SI ≥ 3.

TREATMENT PREPARATION AND ADMINISTRATION:
25 μL of a 1, 5 and 25% concentration of the test substance in DMSO was applied to the dorsal surface of each ear of each mouse, for 3 consecutive days from Day 0. The control group was treated according to the same protocol with the vehicle alone. Local irritation reactions were assessed on Day 1 and 2. In addition, the body weight was measured on Day 1 and 5. On Day 5, 250 μL phosphate buffered saline (PBS) containing 80 μCi/mL of 3H-methyl thymidine (3H-TdR) was injected into the tail vein of each control and treatment mouse. Five hours later, the animals were sacrificed and the draining auricular lymph node of each ear was excised and pooled.The level of 3H-TdR-incorporation was measured in a β-scintillation counter.
The body weight was determined on Day 0 and 5, prior to the treatment.
Positive control substance(s):
not specified
Parameter:
SI
Value:
1.5
Test group / Remarks:
1%
Parameter:
SI
Value:
3.8
Test group / Remarks:
5%
Parameter:
SI
Value:
4.1
Test group / Remarks:
25%
Cellular proliferation data / Observations:
Irritation was not observed and body weight gains were unaffected in all mice. Higher weights of auricular lymph nodes than vehicle control were observed and SI of 25% and 5% groups were higher than 3.

For the control, 1, 5, and 25% concentration groups, the DPM values per lymph node were 1554, 2314, 5887 and 6346, respectively. As the lymph nodes were pooled, these results were determined by dividing the measured value by the number of lymph nodes pooled.

Interpretation of results:
Category 1B (indication of skin sensitising potential) based on GHS criteria
Conclusions:
The simulation index (SI) for the control, 1, 5, and 25% concentration groups was 1.00, 1.5, 3.8 and 4.1, respectively. As the threshold value is 3, the test substance is considered to be a skin sensitiser. The estimated concentration expected to produce a simulation index of 3 (EC3 value) is 3%.
Executive summary:

Skin sensitization potential of test item was evaluated in mice using the Local Lymph Node Assay (LLNA) using method similar to OECD429. CBA/JNCrljmice (three females/group) received the test substance solution (2 5μL/ear)in DMSO at 25%, 5% or 1% on the dorsal surface of each ear.The highest SI (Stimulation Index) of 4.1 was calculated for 25% group and it was concluded that test item is a skin sensitizer under the conditions of this study and the test item was considered to have an EC3 value of 3%.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
From 2014-09-25 to 2014-11-07
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods with acceptable restrictions
Qualifier:
according to guideline
Guideline:
other: GB/T 21608-2008 Chemicals-Test Method of Skin Sensitization
Deviations:
no
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
no
Remarks:
authorized by CNAS
Type of study:
Buehler test
Justification for non-LLNA method:
This in vivo skin sensitisation study was carried out before 10 May 2017, and that in compliance with Chinese GB/T 21608-2008.
Specific details on test material used for the study:
- Purity: 100.0%
- Lot No.: KNC110310
Species:
guinea pig
Strain:
not specified
Sex:
not specified
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Alnimo Laboratory Animal cultivation center, Kunming
- Weight at study initiation: 251- 295 g
- Housing: housed in common environment
- Diet: AIl of the feeds were supplied by Tianqin Biotechnology Ltd., Changsha
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-22
- Humidity (%): 57-67
Route:
epicutaneous, occlusive
Vehicle:
other: 1 % methylcellulose solution
Concentration / amount:
1 g/mL, 0.4 mL
Day(s)/duration:
at 0th, 7th and 14th day, exposure for 6 h
Adequacy of induction:
other: non-irritant substance
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
other: 1 % methylcel1ulose solution
Concentration / amount:
1g/mL, 0.5 mL
Day(s)/duration:
At the 14th day after the latest induction exposure, exposure for 6h
Adequacy of challenge:
highest non-irritant concentration
No.:
#2
Route:
epicutaneous, occlusive
Vehicle:
other: 1 % methylcellulose solution
Concentration / amount:
1g/mL, 0.5 mL
Day(s)/duration:
at 21st days / 6h
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
Treatment group: 20
Negative control group: 10
Details on study design:
RANGE FINDING TESTS:
The highest mild-irritant and nonirritant concentration were determined using 2 Albino Guinea Pig.
MAIN STUDY
- Grouping The negative control group and treatment group were set up in the test, and the number animals for per group were 10 and 20 respectively.
- Dose: According to the result of the preliminary test, the test substance was no irritation. Thereby the concentration of the test substance for induction and challenge was 100%. The volume of the exposure was 0.4 mL and 0.5 mL each animals for induction and challenge respectively. The negative control group was only challenged by test substance.
- Treatment of the test substance: An amount of test item was grinded and then diluted into required concentration 1 g/mL for the test with 1 % methylcel1ulose solution.
- Preparation of test area: Approximately 24 h before the test, about 3 cm x 3 cm areas of fur was removed from both left and right sides of the animals dorsal.
- Exposure:
- Induction exposure: At the 0th, 7thand 14th day, the test substance of 0.4 mL was dropped on the filter paper (2 cm x2 cm) which was applied to the left shaved area, and then covered with 2 ply gauzes and 1 ply paper and held with nonirritating tape for 6 h for each animal in the treatment group. At the end of the exposure period, residual test substance was removed using water.
- Challenge exposure: At the 14th day after the latest induction exposure. The test substance of 0.5 mL was dropped on the filter paper (2 cm x 2 cm) which was applied to the right shaved area, and then covered with 2 ply gauzes and 1 ply paper and held with nonirritating tapes for 6 h for each animal in the negative control and treatment group. After being removed the residual test substance, skin reactions were observed at the 24 and 48 hours. Because of no immune response, rechallenge was done with the same method after 7 days.
- Observation: The health condition and poisoning symptom was observed during the testing period. Skin reactions were graded and recorded at 24 and 48 hours after the challenge and rechallenge exposures.
Challenge controls:
The negative control group was only challenged by test substance.
Positive control substance(s):
no
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
1 g/mL
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
The animals were fine during the testing period
Remarks on result:
no indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
1g/mL
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
The animals were fine during the testing period
Remarks on result:
no indication of skin sensitisation
Reading:
rechallenge
Hours after challenge:
24
Group:
test chemical
Dose level:
1g/mL
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
The animals were fine during the testing period
Remarks on result:
no indication of skin sensitisation
Reading:
rechallenge
Hours after challenge:
48
Group:
test chemical
Dose level:
1 g/mL
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
The animals were fine during the testing period
Remarks on result:
no indication of skin sensitisation
Reading:
other: Challenge and rechallenge
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
The animals were fine during the testing period
Remarks on result:
no indication of skin sensitisation
Remarks:
The results was that of challenge and/or rechallenge of 24 and 48 hours in tanale
Reading:
other: Challenge and rechallenge
Group:
positive control
No. with + reactions:
18
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Remarks:
The results was that of challenge and/or rechallenge of 24 and 48 hours in tanale

Erythema and/or edema were not be found for the negative control and treatment group at 24 and 48 hours, after challenge and rechallenge exposure, the both sensitization rates were 0%.

Interpretation of results:
GHS criteria not met
Remarks:
faint sensitization
Conclusions:
The skin sensitization rate was 0%, and skin sensitization degree of test item was faint sensitization.
Executive summary:

The potential that body produces skin reactions and the sensitization of immunity transfer after the guinea pig exposed repeatedly to test item were determined in accordance with GB/T 21608-2008.

 

Erythema and/or edema were not be found for the negative control and treatment group at 24 and 48 hours, after challenge and rechallenge exposure, the both sensitization rates were 0%.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Skin sensitisation:

in vivo: LLNA, similar to OECD429, SI max: 4.1> 3, EC3 value: 3%, sensitizing

Skin sensitization, similar to OECD406, sensitization rates: 0%; not sensitizing

Respiratory sensitisation: No data available, not minimum required data.

Classification:

Skin: The highest SI (Stimulation Index) was > 3 and EC3 value was 3% in the murine local lymph node assay. Based on criteria from Regulation (EC) 1272/2008 and amendments and Globally Harmonized System Of Classification And Labelling Of Chemicals (GHS) section 3.4.2.2.3, the substance should be classified as category 1B for skin sensitiser.