Cyclohexanecarboxylic acid, 2,2,6-trimethyl-, 2-propen-1-yl ester, (1R,6S)-

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From August 12,2013 to September 3,2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Test conducted according to internationally accepted testing guidelines.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

NACET10419

IUPAC Name:

NACET10419

Constituent 2

Reference substance name:

1450841-11-8

Cas Number:

1450841-11-8

IUPAC Name:

1450841-11-8

Test material form:

other: liquid

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Japan
- Age at study initiation: 8 weeks of age
- Weight at study initiation: 20.3 to 23.8 g.
- Housing: were individually housed in wire mesh metal cages (W 10.0 x D 19.6 x H 13.0 cm). The feeders were exchanged once a week
- Diet: free access to commercial pellet diet, CRF-1
- Water: free access to tap water with an automatic water supply system
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-26 °C
- Humidity (%): 35 to 70 %
- Air changes (per hr): 12 times or more per hour
- Photoperiod (hrs dark / hrs light): 12 hours cycle dark/light (lights on 7 a.m., off 7 p.m.)

Study design: in vivo (non-LLNA)

Positive control substance(s):

yes

Remarks:

α-Hexylcinnamaldehyde

Study design: in vivo (LLNA)

Vehicle:

acetone/olive oil (4:1 v/v)

Remarks:

AOO

Concentration:

Groups Test substance N° of animals
1 vehicle alone 4
2 25 % test substance 4
3 50 % test substance 4
4 100 % test substance 4
5 25 % α-Hexylcinnamaldehyde 4

No. of animals per dose:

4 females x 5 groups (negative control, positive control and 3 doses)

Details on study design:

RANGE FINDING TESTS: A pre-screening test [Exp. No. E93 1 (432-028), non-GLP] was conducted before this main study to determine the concentrations for dosing. In the pre-screening test, three concentrations of 25, 50 and 100% (Acetone and olive oil mixture was used as a solvent, excluding 100 % solution) were selected, and 25 µl each of the dose formulations were applied on the dorsal skin of both auricles of each animal, 2 mice for each concentration, once a day for 3 consecutive days.
The general conditions including the application site condition were observed from 1 to 3 hours after the application on each day of application. As a result, no animals showed any abnormalities. The body weights and ear thickness were measured before the initial application and on Day 6. As a result, none of the animals showed any changes deviating from the criteria of body weights.
From the results mentioned above, for the main study, 100 % was selected as the high concentration because it was expected not to induce any toxic general condition, 25 % or more increase of the ear thickness, dermal erythema with score of 3 or more on the auricles, or more than 5 % of body weight loss, and two lower concentrations of 50 and 25 % (three concentrations in total).

TREATMENT PREPARATION AND ADMINISTRATION: The undiluted solution of the test substance was used for 100 % dose formulation 0.5 ml of the test substance was dissolved in the same volume of acetone and olive oil mixture (AOO) to make the concentration of 50 %. 0.5 ml of this solution was diluted with the same volume of A00 to make the concentration of 25 %.

Positive control substance(s):

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

Statistics:

No statistical analyses were used.

Results and discussion

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

The SI for the 25, 50 and 100 % NACET10419 groups were calculated to be 2.1, 2.6 and 3.0, respectively. Therefore, NACET10419 was judged as positive, and the EC3 value of NACET10419 was calculated as 64% via quadratic regression. On the other hand, DPM/animal of the positive control group was 7800, and the SI of this group was calculated as 9.

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: The DPM/animal of the vehicle control group and 3 test substance groups (25, 50 and 100 %) were 866.7, 2404.8, 2922.5 and 3,420.1, respectively.

Any other information on results incl. tables

No animals showed abnormal clinical signs due to test substance administration during the observation period, no animals showed erythema on the auricles during the observation period.

There were no body weight increases or decreases due to test substance or positive control substance administration during the sensitization period. The weights of lymph nodes in all test substance groups and the positive control group were higher than that in the vehicle control group. No animals showed increases of ear thickness during the sensitization period.

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

NACET10419 was judged to be a sensitizer under the conditions of this study, and was categorized in sub-category 1B of GHS and 1272/2008 classification for skin sensitization.

Executive summary:

A mouse local lymph node assay (LLNA) was conducted to investigate the potential of NACET010419 to induce allergic contact dermatitis. Female CBA/J [SPF] mice were purchased from Charles River Laboratories Japan, Inc., and 20 mice were used for this study. There were 5 dose groups, each consisting of 4 mice, and the animals in each group were exposed to NACET10419 at 1 of the 3 concentration levels, positive control substance or their vehicle (acetone and olive oil mixture: v/v=4: 1).

The results are summarized as follows. The stimulation indices (SI) for the 25, 50 and 100 % NACET10419 groups were calculated to be 2.8, 2.9 and 3.4, respectively. These results were judged to be positive. On the other hand, the SI for the 25% α-Hexylcinnarnaldehyde (HCA, positive control) was calculated to be 9, which was judged to be positive. No animals showed abnormal clinical signs due to test substance administration during the observation period. No animals showed erythema on the auricles during the observation period. The ear thicknesses of all animals were not increased during the sensitizing period. There were no body weight increases or decreases due to test substance administration during the sensitization period. The weights of lymph nodes in all test substance groups and the positive control group were higher than that in the vehicle control group. In conclusion, NACET10419 was judged to be a sensitizer under the conditions of this study, and was categorized in sub-category 1B of GHS and 1272/2008 classification for skin sensitization.