2-Propenoic acid, 2-methyl-, C13-15-branched and linear alkyl esters

Endpoint summary

Description of key information

cLLNA: not sensitzing

Key value for chemical safety assessment

Skin sensitisation

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

2-Propenoic acid, 2-methyl-, C13-15-branched and linear alkyl esters has been previously described to induce an increase of 3H-thymidine (mean SI 6.52), accompanied by a significant increase in ear weight (mean SI 1.28) after treatment with a 25% test-substance preparation in MEK in a LLNA.

The present study, a challenge LLNA, aimed at investigating whether there is a specific secondary immune response and if so at addressing the level required for the induction of a secondary immune response in already sensitized animals.

During the induction treatment groups of 5 female CBA/CaOlaHsd mice each were treated three times with a 25% (w/w) preparation of the test substance in MEK, or with the vehicle alone. During the challenge treatment, the animals were treated once with 2.5%, 5%, 10% and 25% (w/w) preparations of the test substance in MEK or with the vehicle alone. Each animal was treated with 25 μL per ear of the appropriate test-substance preparation or the vehicle applied to the dorsal surfaces of both ears on three consecutive days (day 1, 2 and 3; induction treatment) or on day 21 (challenge treatment). The single application of the 25% test-substance concentration at d21 to the challenge control animals (test group 1) induced statistically significant increases of 3H-thymidine incorporation

into the cells and in the auricular lymph node cell counts, which both failed to reach the cut-off (SI 3 for 3H-thymidine incorporation and SI 1.5 for the auricular lymph node cell counts).

Additionally, statistically significant increase in lymph node weight was observed in the challenge control group. The single application of the 25% test-substance concentration at d21 to the animals previously induced with 25% test-substance concentration (test group 2) elicited a statistically significant increase (compared to the vehicle control group) of 3H-thymidine incorporation into the cells of the auricular lymph nodes, which failed to reach the cut-off. A statistically significant (compared to both the vehicle control and challenge control group) just above the cut-off value for auricular lymph node cell counts was observed. Additonally, a statistically significant (compared to both the vehicle control and challenge control groups) increase of the lymph node weight was noted in this test group. After the single application of 10%, 5% and 2.5% test-substance concentrations at d21 to the animals to the animals previously induced with 25% test-substance concentration (test groups

3, 4 and 5) no biologically relevant increases in 3H-thymidine incorporation, auricular lymph node cell counts, or lymph node weight was observed. The SI’s were statistically significant

(compared to the vehicle control but not the challenge control group) in 3H-thymidine incorporation at 5% and 2.5%, in auricular lymph node cell counts and lymph node weights at 10%, 5% and 2.5%. The test-substance concentrations did not cause an increase (SI ≥ 1.25) in ear weight demonstrating the absence of excessive ear skin irritation. A statistically significant increase was observed in test groups 4 and 5 (compared to both the vehicle control and challenge control groups). 2-Propenoic acid, 2-methyl-, C13-15-branched and linear alkyl esters did not induce specific lymphocyte proliferation in the Challenge Murine Local Lymph Node Assay (cLLNA) under the test conditions chosen.

In the first LLNA study, groups of 5 female CBA/CaOlaHsd mice each were treated with 5%, 10% and 25% (w/w) preparations of the test substance in methyl ethyl ketone (MEK) or with the vehicle alone. No signs of systemic toxicity were noticed in all animals during general observation. When applied as 25% solution in MEK, the test substance induced a biologically relevant (increase to 3-fold or above of control value = stimulation index (SI) ≥ 3), statistically significant and concentration-dependent increase of 3H-thymidine incorporation into the cells from the auricular lymph nodes. The increase at the 10% concentration was statistically significant but failed to reach the cutoff value. Concomitantly, the 25% test-substance preparation induced a biologically relevant, statistically significant and concentration-dependent response (increase to 1.5-fold or above of control value = stimulation index (SI) ≥ 1.5) in the auricular lymph node cell count. The SI of the 10% concentration lies just below the border of biological relevance. In addition, statistically significant increases in lymph node weights were noted at the 10% and 25% concentrations. The 25% concentration caused statistically significant and considerably increased ear weight (SI 1.28) above the cut off Stimulation Index (SI) of 1.25, indicating excessive ear skin irritation. The 5% and 10% concentrations did not cause relevant increases. The increase in ear weight after application of the 25% concentration indicates that the animals of the main study clearly acquire a state of skin irritation, which would be expected to lead to a significantly enhanced response of the lymph nodes after repeated ear treatment. Thus, it remains unclear whether the lymph node responses observed were due to sensitizing properties of the test substance or have to be attributed to cumulated skin irritation effects. 2-Propenoic acid, 2-methyl-, C13-15-branched and linear alkyl esters cannot be conclusively evaluated concerning the skin sensitizing potential in the Local Lymph Node Assay under the test conditions chosen.

Justification for classification or non-classification

Based on the results, the test item was not classified and labelled according to Regulation (EC) No 1272/2008 (CLP).