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Description of key information

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:
10-MAY-2006 TO 24-MAY-2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD guideline study, GLP compliant
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
Test system: Mice, CBA/CaHsdRcc(SPF);
Number of animals for the pre-test: 3 females;
Number of animals for the main study: 16 females;
Number of animals per group: 4 females (nulliparous and non-pregnant);
Age: 8-12 weeks (beginning of acclimatization);
Body weight: 16-24 g;
Identification: Each cage by unique cage card;
Randomization: Randomly selected by computer algorithm at time of delivery.
Acclimatization: Under laboratory conditions after health examination. Only healthy animals were used for the study.

Conditions
Standard Laboratory Conditions.
Air-conditioned with ranges for room temperature of 22 ± 3 °C and relative humidity 30-70 % and approximately 10-15 air changes per hour. There was a 12 hours artificial fluorescent light/12 hours dark, music during the light period.
Accommodation: Individual in Makrolon type-2 cages with standard softwood bedding.
Diet: Pelleted standard Provimi Kliba 3433 mouse maintenance diet (batch 001/06) ad libitum.
Water: Community tap water, ad libitum.
Vehicle:
dimethylformamide
Concentration:
2.5%, 5% and 10% in 25 µl of DMF
No. of animals per dose:
4
Details on study design:
Three groups each of four female mice were treated daily with the test item at concentration of 2.5%, 5% and 10% in DMF by topical application to the dorsum of each ear lobe for three consecutive days. Five days after the first topical application, the mice were injected intravenously into a tail vein with radiolabelled thymidine(3H-methyl thymidine, 3HTdR). Approximately five hours after intravenous injection, the mice were sacrificed, the draining auricular lymph nodes excised and pooled per group. Single cell suspensions of lymph node cells were prepared from pooled lymph nodes which were subsequently washed and incubated with trichloroacetic acid overnight. The proliferative capacity of pooled lymph node cells was determined by the incorporation of 3HTdR measured in a ß–scintillation counter.
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.
Positive control results:
S.I. of 1.8, 2.9 and 6.2 of determined with alpha-hexylcinnamaldehydewere at concentrations of 5%, 10% and 25%, respectively, in acetone: olive oil, 4:1 (v/v). Alpha-hexylcinnamaldehyde was therefore found to be a skin sensitizer in the LLNA tests and an EC3 value of 10.5% was derived.
Parameter:
SI
Remarks on result:
other: S.I. of 10.9, 16.8 and 23.0 were determined with the test item at concentrations of 2.5%, 5% and 10%, respectively, in DMF.
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: DPM of 9983, 15302 and 20976 were determined with the test item at concentrations of 2.5%, 5% and 10%, respectively, in DMF.
Interpretation of results:
sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
S.I. of 10.9, 16.8 and 23.0 were determined with the test item at concentrations of 2.5%, 5% and 10%, respectively, in DMF and the test item was therefore found to be a potential skin sensitizer in the LLNA tests.
Executive summary:

Three groups each of four female mice were treated daily with the test item at concentration of 2.5%, 5% and 10% in DMF by topical application to the dorsum of each ear lobe for three consecutive days. Five days after the first topical application, the mice were injected intravenously into a tail vein with radiolabelled thymidine (3H-methyl thymidine, 3HTdR). Approximately five hours after intravenous injection, the mice were sacrificed, the draining auricular lymph nodes excised and pooled per group. Single cell suspensions of lymph node cells were prepared from pooled lymph nodes which were subsequently washed and incubated with trichloroacetic acid overnight. The proliferative capacity of pooled lymph node cells was determined by the incorporation of 3HTdR measured in a ß–scintillation counter.

All treated animals survived the scheduled study period.

No clinical signs were observed.

After the first topical application, the dark red stain was found at both dosing sites in all mice of the test item group, persisting for the remainder of the in-life phase of the study.

 

In the study, S.I. of 10.9, 16.8 and 23.0 were determined with the test item at concentrations of 2.5%, 5% and 10%, respectively, in DMF. The EC3 value cannot be determined, since this calculation requires a S.I. values less than 3. Although according to the data reported above, the EC3 can be derived to be < 1%, thus the substances shows a high sensitizing potential.

Based on these results, the substance is to be classified as Skin Sensitizer category 1A according to the CLP regulation (Regulation EC No. 1272/2008 as amended).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)
Additional information:

This skin sensitization study was performed according to OECD Guideline 429 under GLP compliance. Three groups each of four female mice were treated daily with the test item at concentration of 2.5 %, 5 % and 10 % in DMF by topical application to the dorsum of each ear lobe for three consecutive days. All treated animals survived the scheduled study period. No clinical signs were observed. After the first topical application, the dark red stain was found at both dosing sites in all mice of the test item groups., persisting for the remainder of the in-life phase of the study. In the study, S.I. of 10.9, 16.8 and 23.0 were determined with the test item at concentrations of 2.5 %, 5 % and 10 %, respectively, in DMF. The test item was therefore found to be a potential skin sensitizer in the LLNA test.


Migrated from Short description of key information:
The test item was found to be a potential skin sensitizer in the LLNA tests.

Justification for selection of skin sensitisation endpoint:
Guideline study with GLP

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The test substance shall be classified as Category 1A according to the CLP regulation (Regulation EC No. 1272/2008) and as skin sensitiser Xi, R43 according to DSD (Directive 67/548/EEC). Data on respiratory irritation are lacking.