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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1998 and 1999
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: A published study conducted following a protocol which was the basis for establishing the corresponding guideline adopted in 2002.
Justification for type of information:
Information used for read across to Acalea.
Cross-reference
Reason / purpose:
read-across: supporting information
Reference
Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Results derived from a valid read across, with adequate and reliable documentation / justification.
Justification for type of information:
The read across justification is presented in the endpoint summary. The corresponding documentation file is also attached there.
Reason / purpose:
read-across source
Key result
Parameter:
EC3
Remarks:
(%)
Value:
6.6
Remarks on result:
other: Based on read across information.
Parameter:
SI
Value:
1.7
Remarks on result:
other: Dose 2.5%
Parameter:
SI
Value:
2.1
Remarks on result:
other: Dose 5%
Parameter:
SI
Value:
4.4
Remarks on result:
other: Dose 10%
Parameter:
SI
Value:
8.1
Remarks on result:
other: Dose 25%
Parameter:
SI
Value:
14.5
Remarks on result:
other: Dose 50%
Interpretation of results:
other: Skin sensitiser, Category 1B
Remarks:
according to EU CLP Regulation (EC) No. 1272/2008 and its amendments.
Conclusions:
For Acalea an EC3 of 6.6% is derived for skin sensitization based on read across information from Hexyl Cinnamic Aldehyde.

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
A comparison of statistical approaches to the derivation of EC3 values from local lymph node assay dose responses
Author:
Basketter DA, Lea LJ, Dickens A, Briggs D, Pate I, Dearman RJ, Kimber I
Year:
1999
Bibliographic source:
Journal of Applied Toxicology 19, 261-266
Reference Type:
publication
Title:
Temporal stability of local lymph node assay responses to hexyl cinnamic aldehyde
Author:
Dearman RJ, Hilton J, Evans P, Harvey P, Basketter DA, Kimber I
Year:
1998
Bibliographic source:
Journal of Applied Toxicology 18, 281-284

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
GLP compliance:
not specified
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Test material form:
liquid

In vivo test system

Test animals

Species:
mouse
Strain:
other: CBA/Ca
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan UK, Bicester, UK
- Age at study initiation: 8-16 weeks
- Weight at study initiation:no data
- Housing: in groups of 4 in metal cages on a flushing mouse rack
- Diet (e.g. ad libitum): Ad libitum; SDS PCD pelleted diet, Special Diets Services Ltd.
- Water (e.g. ad libitum): Ad libitum
- Acclimation period:no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C):21+/-2
- Humidity (%):55+/-10
- Air changes (per hr):no data
- Photoperiod (hrs dark / hrs light):12/12

IN-LIFE DATES: From: To:no data

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
0, 2.5, 5.0, 10.0, 25.0, 50.0% w/v
Solutions were prepared freshly before each application
No. of animals per dose:
4
Details on study design:
RANGE FINDING TESTS: none conducted.

MAIN STUDY Data is provided from 5 separate tests conducted in the same test laboratory within the period of 10 months.
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method:pooled
- Criteria used to consider a positive response: SI>=3 (linear interpolation)

TREATMENT PREPARATION AND ADMINISTRATION:
See further details under 'Any other information on materials and methods inc. tables'
The method used was that described by:
Kimber and Basketter. The murine local lymph node assay; collaborative studies and new directions: a commentary. Food Chem. Toxicol. 30, 165-169 (1992).
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
Three statistical models were used to derive the EC3 from the data: Linear interpolation, Quadratic regression, Richard's model. They were performed separately for each experiment/test.
Analysis of covariance was used to compare stimulation indices across experiments.

Results and discussion

Positive control results:
Results for hexylcinnamic aldehyde are provided below.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Value:
1.7
Remarks on result:
other: Test 1: Dose 2.5% w/v
Parameter:
SI
Value:
2.1
Remarks on result:
other: Test 1: Dose 5% w/v
Parameter:
SI
Value:
4.4
Remarks on result:
other: Test 1: Dose 10% w/v
Parameter:
SI
Value:
8.1
Remarks on result:
other: Test 1: Dose 25% w/v
Parameter:
SI
Value:
14.5
Remarks on result:
other: Test 1: Dose 50% w/v
Parameter:
SI
Test group / Remarks:
Results of 5 tests.
Remarks on result:
other: Doses: 0, 2.5, 5, 10, 25, 50% w/v Test 1: 1, 1.7, 2.1, 4.4, 8.1, 14.5 Test 2: 1, 1.7, 2.1, 2.4, 7.2, 14.1 Test 3: 1, 1.3, 2.1, 2.7, 7.8, 13.4 Test 4: 1, 2.2, 3.2, 7.1, 13.9, 17.6 Test 5: 1, 1.0, 1.4, 2.0, 8.7, 11.6
Parameter:
other: disintegrations per minute (DPM)
Test group / Remarks:
Results of 5 tests.
Remarks on result:
other: see Remark
Remarks:
5 different tests conducted: Test 1: DPM/node - 472, 786, 1002, 2054, 3841, 6844 Test 2: DPM/node - 475, 817, 998, 1127, 3396, 6698 Test 3: DPM/node - 325, 437, 685, 876, 2539, 4356 Test 4: DPM/node - 159, 355, 508, 1124, 2207, 2795 Test 5: DPM/node - 188, 192, 256, 371, 1639, 2186

Any other information on results incl. tables

There was a statistically significant (p<0.01) difference between the inter-experimental SI. This is because a higher than average SI was determined in Test 4. There was also a steeper shape in the dose-response curve for the Test no. 4 data making it a p<0.01 for the quadratic component. Both significant differences were lost when data from Test 4 were excluded from the analysis. The results from Test 4 are considered be outliers.

Table 1. EC values derived:

Test no

Linear EC3

Quadratic EC3

Richard’s EC3

1

6.6

6.9

6.9

2

11.3

9.6

11.1

3

10.6

8.7

10.4

4

4.4

4.0

3.9

5

11.5

9.2

13.1

Applicant's summary and conclusion

Interpretation of results:
sensitising
Conclusions:
A EC3 between 6.6-11.5% can be derived from four different tests conducted, using three different calculation methods
Executive summary:

Hexyl cinnamaldehyde is recommended as a positive control by the OECD guideline 406 for Skin Sensitisation. As such, the response of hexyl cinnamaldehyde in the Local Lymph Node Assay from five different experiments conducted in a single laboratory over a 10 month period was analysed. Hexylcinnamaldehyde in 0, 2.5, 5, 10, 25, 50% w/v in 4:1 acetone/olive oil was administered onto the dorsal ears of mice following a protocol equivalent to the OECD guideline 429 Skin Sensitisation: Local Lymph Node Assay adopted later. Hexyl cinnamaldehyde induced a positive response in all five tests (SI>3). The pooled stimulation indices were used to calculate the EC3 values using three different statistical approaches (Linear interpolation, Quadratic regression and Richard's model) which resulted in comparable EC3 values. Excluding the results of one of the tests which was considered to be an outlier, the EC3 range for hexylcinnamaldehyde was 6.6 -11.5% under the conditions of these experiments.