Registration Dossier

Administrative data

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:
other: Guideline study and GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymphnode assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
GRO 266
SAT 070058
Batch-No.: GRO-RN-9547-185

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female

Study design: in vivo (LLNA)

Vehicle:
other: Ethanol:Elix water (7:3)
Concentration:
10, 25, 50 %
No. of animals per dose:
5
Details on study design:
Vehicle
Ethanol/Elix water (7:3 v/v)
Preliminary irritation study
A preliminary iritation study was conducted in order to select the highest test substance concentration to be used in the main study. In principle,
this concentration should be well tolerated systemically by the animals and may give moderate irritation (grade 2) at the highest.
A series of two test substance concentrations was tested, selected from the series: 100 % (undiluted), 50%, 25%, 10%, 5%, 2.5%, 1% and if needed further lower concentrations using the same steps. The highest concentration, selected from this series, was the maximum concentration that be technically be applied.
The test system, procedures and techniques were identical to those used during Days 1 - 3 of the main study unless otherwise specified. Two you adult animals were selected (8-14 weeks old). Each animal was treated with one concentration on three consecutive days. Approximately 3-4 hours after the last exposure, the ear was cleaned of residual test substance with tap water and the irritation was assessed. Bodyweights were determined on Day 3. The animals were sacrificed after the final observatio and no necropsy was performed.
Main study
Three groups of five animals were treated with one test substance concentration per group. The highest test substance concentration was selected from the preliminary irritation study. One group of five animals was treated with vehicle.
Induction - Days 1, 2, 3
The dorsal surface of both ears was epidermally treated (25 µl/ear) with the test substance concentration, at approximately the sam time per day. The concentrations were mixed thoroughly using a vortex mixer immediately prior to dosing. The control animals were treated the same as the experimental animals, expected that, instead of the test substance, the vehicle alone was administered.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: SI: 10 % = 1.0 25 % = 2.3 50 % = 1.3 vehicle = 1.0
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: DPM +- SEM 10% = 142 +- 21 25% = 334 +- 61 50% = 181 +- 45 0 % (vehicle) = 143 +- 52

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information
Conclusions:
The SI values calculated for the substance concentrations 10, 25, and 50 % were 1.0, 2.3 and 1.3 respectively.
Since there was no indication that the test substance elicit an SI > 3 when tested up to 50 %, GRO 266 was considered to be a non skin sensitizer.
The six monthly reliability check with Hexylcinnamaldehyde, indices that the Local Lymph Node Assay as performed at NOTOX is an appropriate model for testing for contact hypersensitivity.
Executive summary:

Assessment for Contact Hypersensitivity to GRO 266 in the Mouse (Local Lymph Node Assay),

The study was carried out based on the guidelines described in:

OECD, Section 4, Health Effects, No. 429 (2002),

EC, Council Directive 67/548/EEC, Annex V, B.42 (2004);

EPA, OPPTS 870.2600 (2003) "Skin Sensitisation".

Test Substance concentrations selected for the main study were based on the results of a preliminary study.

In the main study, three groups of five experimental animals were treated with test substance concentration of 10%, 25% and 50% on three consecutive days, by open application on the ears. Five vehicle control animals were similarly treated, but with vehicle alone (Ethanol/water). Three days after the last exposure, all animals were injected with 3H-methyl thymidine and after five hours the draining (auricular) lymph node cells excised.

After precipitating the DNA of the lymph node cells, radioactivity measurements were performed. The activity was expressed as the number of Disintegrations Per inute (DPM) and a stimulation index (SI) was subsequently calculated for each group.

Slight irritation was observed in one ear of one animal at 50%. No oedema was observed in any of the animals examined. The irritation of the ears as shown by one animal was considered not to have a toxicologically significant effect on the activity of the nodes.

The majority of nodes were considered normal in size, expect for the nodes of one animal treated at 10 %, two animals treated at 25 % and one animal at 50 %. No macrosopic abnormalities of the aurrounding area were noted.

Mean DPM/animal values for the experimenta groups treated with substance concentrations 10, 25 and 50% were 142, 334 and 181 respectively. The mean DPM/animal value for the vehicle control group was 143.

The SI values calculated for the substance concentrations 10, 25 and 50 % were 1.0, 2.3 and 1.3 respectively.

Since there was no indicated thet the test substance elicit an SI>3 when tested up to 50%, GRO 266 was considered to be a non skin sensitizer.

No ECETOC classification is applicable.