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Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Administrative data

skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD guideline, GLP study

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
GLP compliance:
yes (incl. QA statement)
Type of study:
mouse local lymph node assay (LLNA)

Test material

Constituent 1
Reference substance name:
LCE 10046
LCE 10046
Details on test material:
Test Substance/Itern : LCE 10046
Chemical name : Not applicable
Manufactured date : 31 .08.2009
Retest date : 31.08.2011
Purity (as per Acid value :1 .8 mg KO1-1/g
Certificate of Hydroxyl value: 439 mg KOH/g
Physical appearance : Pearls appearance
Storage condition : Ambient (+18 to +36°C)
Note: Date of receipt of test item at test facility: 17.06.20 10 Test item code by test facility: S015-02

In vivo test system

Test animals

Details on test animals and environmental conditions:
Species : Mice
Strain : CBA/Ca
Source : Harlan, The Netherlands
No. of groups : 7 groups
Vehicle control (G1)
Positive control (G2)
Test item: Five concentrations (G3 to 07)
Age at treatment : 11-12 weeks
Body weight range : 20.14 to 24.06 grams
Identification : Identification of individual animals: by cage card and body markings using turmeric solution.
Acclimatization : After physical examination for good health and the suitability for the study, the animals were acclimatized for 6 days before start of the treatment. Animals were nuliiparous and nonpregnant.

Room Number: Room No. SC-4
Mice were housed in an environment controlled room at 21 to 23°C and relative humidity of 65 to 67 per cent. The photoperiod was 12 hours light and 12 hours darkness, light hours being 06.00 to 18.00 hours approximately. Adequate fresh air suppiy of 14.8 air changes/hour was maintained in the experimental room.
The maximum and minimum temperature and relative humidity in the experimental room were recorded once daily. The relative humidity in the experimental room was calculated daily from dry and wet bulb temperature recordings.

Animals were housed individually in solid floor standard polysulfone cages (Size: Approximately L 360 x B 205 x H 140 mm), with stainless steel top grill having facilities providing pelletted food and drinking water in polycarbonate bottle with stainless steel sipper tubes. Steam sterilized clean paddy husk was used as bedding and changed along with the cage twice a week. Cages were placed on 5 tier racks.

Diet and Water
The experimentai animals were provided ad libitum standard pelletted food (Ssniff mice peliet food — maintenance manufactured by Ssniff Spezialditen GmbH., Ferdinand-Gabriel-Weg 16, D-59494 Sôest, Germany) and deep bore-well water passed through activated charcoal filter and exposed to UV rays in Aquaguard on-une water filter-cum-purifier (manufactured by Eureka Forbes Ltd., Mumbai -400001, India).
The food and water provided to the animals was tested for contaminants.

Study design: in vivo (LLNA)

acetone/olive oil (4:1 v/v)
positive control: 30% v/v

test material: 1 / 5 / 10 / 25 / 50% w/v
No. of animals per dose:
Details on study design:
Animals were randomly distributed to different groups by body weight stratification method. The mice procured for the study were weighed and grouped into body weight ranges. These body weight stratified animals were distributed to au the study groups. Mice with extreme body weights were removed from the study.

Dose Preparation and Analysis
Concentrations tested for the irritancy screen were selected based upon miscibility/solubility in an appropriate LLNA vehicle. The toxicity data regarding the irritation potential was also taken into consideration.
The required quantity of the test item was rnelted in hot water bath (60 - 70 °C). The test item LCE 10046 was mixed with AOO to obtain concentrations of 1, 5, 10,25 and 50 w/v, vortexed and kept in water bath until application. The test item solutions were preparcd daily just prior to dosing. Preparation of the dosing materials was documented in the study file. The concentration of the dosing solution was not verified analytically.

Irritation Screen
Prior to the LLNA main study, concentrations of 1, 5, 10, 20, 35 and 50% w/v of LCE 10046, suspended in AOO were evaluated for irritation potential as measured by erythema of the ears.
Both ears of six female mice (one mouse/concentration) were topically treated once daily for three consecutive days with one of the concentrations of the test item.
The test item was administered using an adjustable micropipette with disposable tips. All mice received 25 uL (12 and 13 uL to each ear) of one concentration of the test item, spread over the dorsal surface of each ear in a manner to prevent test item loss. Similarly the vehicle was applied to the ears of one animal. Both ears were observed prior to application of the test item, and erythema was valuated on Days 2, 3 and 6. All mice were weighed on Days 1 and 6. Erythema scores and body weight data following the test item applications were compared to the response ofthe animals treated with vehicle alone (irritation to mouse ears is only relevant in the context of the LLNA and should not be interpreted as an indicator for an irritation potential in humans).

Dermal Sensitization
The application of the test item (25 uL/ear) was made on the dorsum of both ears as described above. Five female mice/group received the vehicle (AOO), or the positive control substance (30% a-hexylcinnamaldehyde), or 1, 5, 10,25 and 50 w/v of LCE 10046, once daily for three consecutive days. Ears were inspected prior to each application of the test item solution, and erythema was evaluated on days 2, 3 and 6. Ail mice were weighed on days I and 6.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:
The sensitivity of this LLNA test was demonstrated via the response from the positive control (30% HCA in AOO), which elicited a stimulation index (SI) of 5.36, in comparison with the vehicle-treated mice.

In vivo (LLNA)

Resultsopen allclose all
Remarks on result:
other: negative control group: 1 positive control group: 5.36 1% group: 1.4 5% group: 1.28 10% group: 1.04 25% group: 1.46 50% group: 1.38
other: disintegrations per minute (DPM)
Remarks on result:
other: Average per group vehicle group: 1351.2 positive control group: 7239.4 1% group: 1884.2 5%: 1732.4 10%: 1406.2 25%: 1967.4 50%: 1863.4

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Migrated information
A topical application with 1, 5, 10, 25 and 50% w/v LCE 10046 elicited a stimulation index (SI) of 1.40, 1.28, 1.04, 1.46 and 1.38, respectively. The test item LCE 10046 did not elicit a stimulation index (SI) that met the 3X threshold, thus indicating a lack of dermal sensitization potential in the mouse
Executive summary:

This Local Lymph Node Assay (LLNA) was conducted to evaluate the potential of LCE 10046 to cause contact sensitization by measuring lymphocyte proliferative response from auricular lymph nodes following topical application of the test item to the female CBA/Ca mouse ear.

Screening Study : Three daily topical application at 1, 5, 10, 20, 35 and 50% w/v of LCE 10046 in acetone:olive ou [AOO] (4:1 mixture v/v) were given to one animal at each dose level. There were no clinical signs of toxicity, no effect on body weight and no reactions at site of application. Results from this study were used to determine the dosing concentration for the main study.

LLNA main study : Five female CBA/Ca mice/group received the vehicle (AOO) or 30% a-hexylcinnamaldehyde (HCA: positive control in AOO) or 1, 5, 10, 25 and 50% wlv of LCE 10046 in AOO on days 1 to 3. On day 6, uptake of 3H-methyl thymidine into the auricular lymph nodes draining the site of test item application was measured five hours post administration. Proper conduct of the LLNA was confirmed via a positive response using 30% a-hexylcinnamaldehyde, a contact sensitizer, which elicited proliferation with a Stimulation Index (SI) value of 5.36, in comparison to vehicle-treated mice.

The test item LCE 10046 did not elicit erythema at the tested concentrations and no significant effect on body weight gain.

The test item LCE 10046 at dose concentrations of 1, 5, 10, 25 and 50% w/v elicited proliferative response with SI of 1.40, 1.28, 1.04, 1.46 and 1.38, respectively in comparison with the vehicle-treated mice.

LCE 10046 did not demonstrate dermal sensitization potential in the mouse LLNA, as the lymph nodes draining the area of topical application did not elicit a proliferative response greater than the 3X threshold.