3,5,8,10,13,15,18,20,23,25,28,30,33,35,38,40,41,43,45,47,51,53,55,57,59,61,63, 65,67,69,71,73-dotriacontazapentacosacyclo [35.3.3.36,7.311, 12.316,17.321,22.326,27.331,32.22,41.236,43.13,40.15,8.110,13.115,18.120,23.125,28.130,33.135,38.145,51.147,73.153,55.157,59.161,63.165,67.169,71] octacontane-44,46,48,49,50,52,54,56,58,60,62,64,66,68,70,72-hexadecone, hydrochloride hydrate

Endpoint summary

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

OGYI/38593-5/2012

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

other: CBA/J Rj

Sex:

female

Vehicle:

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

Concentration:

25% CUCURBIT[8]URIL in Acetone : Olive oil 4:1 (v/v) mixture
10% CUCURBIT[8]URIL in Acetone : Olive oil 4:1 (v/v) mixture
5% CUCURBIT[8]URIL in Acetone : Olive oil 4:1 (v/v) mixture
Positive control - 25% (w/v) a-Hexylcinnamaldehyde (dissolved in AOO).
Negative control - Acetone : Olive oil 4:1 (v/v) mixture

No. of animals per dose:

Four mice per group.

Positive control substance(s):

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

Key result

Parameter:

SI

Value:

1.1

Test group / Remarks:

5%

Key result

Parameter:

SI

Value:

1.3

Test group / Remarks:

10%

Key result

Parameter:

SI

Value:

1

Test group / Remarks:

25% (w/v)

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: see Remark

Remarks:

DPM was measured for each pooled group of nodes. The measured DPM values were corrected with the background DPM value (“DPM”). The average of the two measured DPM values of 5% (w/v) TCA solutions was used as background DPM value. The results were expressed as “DPN” (DPM divided by the number of lymph nodes) following the industry standard for data presentation. The DPM values were 2634.5, 3407.5 and 2983.5 at concentrations of 25, 10 and 5% (w/v), respectively.

Individual Body Weights for all Animals with Group Means

Identity Number	Animal Number	Test group name	Initial body weight (g)	Terminal body weight (g)	Change (%)^#
7467	1	Negative vehicle control (AOO)	22.0	22.1	0.5
7539	2		21.5	21.5	0.0
7484	3		21.9	22.4	2.3
7508	4		21.6	22.3	3.2
		Mean	21.8	22.1	1.5
7469	5	CUCURBIT[8]URIL 25% (w/v) in AOO	22.1	21.6	-2.3
7546	6		21.7	21.0	-3.2
7490	7		21.7	22.5	3.7
7516	8		21.9	21.7	-0.9
		Mean	21.9	21.7	-0.7
7475	9	CUCURBIT[8]URIL 10% (w/v) in AOO	22.0	21.3	-3.2
7550	10		21.7	22.9	5.5
7494	11		21.6	22.3	3.2
7525	12		21.9	22.2	1.4
		Mean	21.8	22.2	1.7
7480	13	CUCURBIT[8]URIL 5% (w/v) in AOO	22.1	23.3	5.4
7554	14		21.9	20.7	-5.5
7498	15		21.5	22.6	5.1
7532	16		21.7	21.7	0.0
		Mean	21.8	22.1	1.3
7472	17	Positive control (25% (w/v) HCA in AOO)	22.0	22.4	1.8
7493	18		21.9	22.3	1.8
7512	19		21.7	23.0	6.0
7537	20		21.5	22.2	3.3
		Mean	21.8	22.5	3.2

Notes:

*: Terminal body weights were measured on Day 6.

#: = (Terminal Body Weight – Initial Body Weight) / Initial Body Weight x 100

DPM, DPN and Stimulation Index Values for all Groups

Test Group Name	Measured DPM / group	DPM	Number of Lymph Nodes	DPN	Stimulation Index
Background (5% (w/v) TCA)	31 28	-	-	-	-
Negative vehicle control (AOO)	2638	2608.5	8	326.1	1.0
CUCURBIT[8]URIL 25% (w/v) in AOO	2664	2634.5	8	329.3	1.0
CUCURBIT[8]URIL 10% (w/v) in AOO	3437	3407.5	8	425.9	1.3
CUCURBIT[8]URIL 5% (w/v) in AOO	3013	2983.5	8	372.9	1.1
Positive control (25% (w/v) HCA in AOO)	23468	23438.5	8	2929.8	9.0

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

In conclusion, under the conditions of the present assay, CUCURBIT[8]URIL, tested in a suitable vehicle, was shown to have no sensitisation potential
(non-sensitizer) in the Local Lymph Node Assay.

Executive summary:

The purpose of this study was to determine the skin sensitisation potential of CUCURBIT[8]URIL test item following dermal exposure in the Local Lymph Node Assay. Study performed to OECD Guidelines for Testing of Chemicals No. 429 “Skin Sensitisation: Local Lymph Node Assay”(22 July 2010) Commission Regulation (EC) No 440/2008 of 30 May 2008, B.42. “Skin Sensitisation: Local Lymph Node Assay” (Official Journal L 142, 31/05/2008) as amended by Commission Regulation (EU) No 640/2012 of 6 July 2012 (Official Journal L 193, 20/07/2012).

In conclusion, under the conditions of the present assay, CUCURBIT[8]URIL, tested in a suitable vehicle, was shown to have no sensitisation potential (non-sensitizer) in the Local Lymph Node Assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

The aim of this study was to determine the skin sensitisation potential of CUCURBIT[8]URIL following dermal exposure. The study was performed with vertebrate animals as no regulatory in vitro alternative is available. The minimum number of animals was used, corresponding to the regulatory guidelines being followed.

Based on the results of the preliminary solubility / compatibility test, preliminary toxicity tests and on the recommendations of the OECD Guideline, the test item was formulated in Acetone : Olive oil 4:1 (v/v) mixture (abbreviated as AOO). Due to the physical and chemical characteristics of the test item, the highest achievable concentration was 25% (w/v).

The Preliminary Irritation / Toxicity Test was performed in CBA/J Rj mice using two doses (25 and 10% (w/v) in AOO). Based on the observations recorded in the preliminary test, 25% (w/v) was selected as top dose for the main test.

In the main assay, twenty female CBA/J Rj mice were allocated to five groups of four animals each:

-three groups received CUCURBIT[8]URIL (formulated in AOO) at 25, 10 and 5% (w/v) concentrations,

-the negative control group received the vehicle (AOO),

-the positive control group received 25% (w/v) HCA (dissolved in AOO).

The test item solutions were applied on the dorsal surface of ears of experimental animals (25 μL/ear) for three consecutive days (Days 1, 2 and 3). There was no treatment on Days 4, 5 and 6. On Day 6, the cell proliferation in the local lymph nodes was measured by incorporation of tritiated methyl thymidine (3HTdR) and the values obtained were used to calculate stimulation indices (SI).

No mortality or signs of systemic toxicity were observed during the study. No treatment related body weight loss was observed in the test item treated animals. Minimal amount of test item precipitate was observed on the ears of the animals in the 25% (w/v) test item treated group on Days 1-3. There were no indications of any irritancy at the site of application.

The stimulation index values were 1.0, 1.3 and 1.1 at concentrations of 25, 10 and 5% (w/v), respectively.

The result of the positive control substance a-Hexylcinnamaldehyde (HCA) dissolved in the same vehicle was used to demonstrate the appropriate performance of the assay. A significant lymphoproliferative response (stimulation index value of 9.0) was noted for the positive control chemical, this result confirmed the validity of the assay.

In conclusion, under the conditions of the present assay, CUCURBIT[8]URIL, tested in a suitable vehicle, was shown to have no sensitisation potential (non-sensitizer) in the Local Lymph Node Assay.

Migrated from Short description of key information:
Not sensitising to mice in the local lymph node assay (LLNA).

Justification for selection of skin sensitisation endpoint:
Endpoint conclusion taken from experimental study using OECD Guideline 429 and EU Method B42 in accordance with GLP.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

The substance is not classified under CLP as it does not fill the requirements for skin sensitisation classification.