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:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

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 lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
Test item: Levodione
CAS number: 21800-83-9
Batch number: 242027N
Physical state: Solid, crystalline powder
Colour: White
Storage: At a temperature of 15 to 30 °C

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
Species and strain: CBA/Ca Ola Hsd mice
Source: TOXI-COOP ZRT.
Hygienic level at arrival: SPF
Hygienic level during the study: Good conventional
Number of animals: 24 animals/main test (4 animals/treatment group)
Sex: Female, nulliparous, non-pregnant
Age of animals: Young adult mice; 10-11 weeks old (at start of the main test)
Body weight range at starting: 18.0-23.6 g
Acclimatization time: 14 days
Animal health: Only healthy animals were used

Housing during acclimatization period: Grouped caging in small groups
Housing during the test: Grouped caging (4 animals/cage)
Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 22 ± 3 °C
Relative humidity: 30 – 70 %

Study design: in vivo (LLNA)

Vehicle:
dimethylformamide
Remarks:
AOO (acetone:olive oil, 4:1) for positive control groups
Concentration:
The test item was administered at three different concentrations (5; 2,5; 1 w/v%) according to the results of the dose range finding test.
No. of animals per dose:
4 animal/dose
Details on study design:
Selection of test item concentrations was based on the results of a formulation evaluation and also results of preliminary irritation/toxicity test to find an appropriate solvent and the maximum applicable concentration according to the relevant guidelines. Based on the preliminary test results the test item was examined in the main test at 5 %, 2.5 % and 1 % (w/v) concentrations as formulations in the selected vehicle of N,N-Dimethylformamide (DMF). Appropriate positive control (α-Hexylcinnamaldehyde [HCA] in 25 % (w/v) concentration), furthermore two negative control groups dosed with the vehicles of the test and positive control groups, respectively, were employed.
The positive control item (25 % (w/v) HCA in Acetone: Olive oil 4:1 (v/v) mixture, AOO) induced the appropriate stimulation over the control, thus confirming the validity of the assay.
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: see Remark
Remarks:
The stimulation index (SI = the DPN of a treated (positive control or test item) group divided by the DPN of the respective negative control group) for each treatment group was also calculated. A stimulation index of 3 or greater is an indication of a positive result. When monotonic dose-response is seen, attempt is made to calculate EC3 (dose calculated to induce a stimulation index of 3) with the help of linear regression on dose-response relationship. All calculations were made by Microsoft Excel Software. (See Table1 for the values.)
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: see Remark
Remarks:
DPM (disintegration per minute) was measured for each treatment group. The measured DPM values were corrected with the background DPM value. The average of the two measured DPM values of 5 % (w/v) TCA solutions was used as the background DPM value. The results were expressed as DPN (DPM divided by the number of pooled lymph nodes). (See Table1 for the values.)
Parameter:
SI
Value:
1.9
Remarks on result:
other: Levodione 5 % in DMF
Parameter:
SI
Value:
1.6
Remarks on result:
other: Levodione 2.5 % in DMF
Parameter:
SI
Value:
2.8
Remarks on result:
other: Levodione 1 % in DMF

Any other information on results incl. tables

Test Group Name

Measured

DPM/group

Group*

DPM

DPN

(DPM/Node)

Stimulation

Index Values

Vehicle control for the positive control: AOO

18071

18050.0

2256.3

1.0

Positive control:
25 % HCA
in AOO

86012

85991.0

10748.9

4.8

Levodione
5 %
in DMF

7552

7531.0

941.4

1.9

Levodione
2.5
% in DMF

6409

6388.0

798.5

1.6

Levodione
1 %
in DMF

11056

11035.0

1379.4

2.8

Vehicle control for the test item: DMF

4019

3998.0

499.8

1.0

 *Group DPM = measured DPMgroup- average DPMbackground

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Remarks:
Migrated information
Conclusions:
In conclusion, under the conditions of the present Local Lymph Node Assay, Levodione (CAS No. 21800-83-9) tested at the maximum attainable concentration of 5 % (based on solubility) and at concentrations of 2.5 % and 1 % (w/v) as formulations in an appropriate vehicle (DMF) was shown to have no skin sensitization potential.