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: Well documented study according to internationally accepted guidelines and CLP compliant.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report Date:
2013

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: crystalline
Details on test material:
Test item: IMIDAZOLE ETHANOL (CAS No. 24155-42-8)
Batch No.: L36066N
Physical state: Light-yellow powder
Active ingredient content (titrimetry): 92.1%
Impurities (HPLC): 9.18%
Storage: 15-30°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 mice
Source: TOXI-COOP ZRT.
Hygienic level at arrival: SPF
Hygienic level during the test: Good conventional
Number of animals: 4 animals/preliminary test (2 animals/treatment group)
Sex: Female, nulliparous, non pregnant
Age of animals: Young adult mice; 11-12 weeks old (at start of the preliminary test)
Body weight range at starting: 17.0-22.3 g
Acclimatization time: 7 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 four different concentrations (25; 10; 5; 2.5 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 the best solvent and the maximum applicable concentration according to the relevant guidelines. The test item was tested at the solubility limit of 25 % and at three lower concentrations (10 %, 5 % and 2.5 %) in N,N-Dimethylformamide (DMF). Appropriate positive control (α-Hexylcinnamaldehyde [HCA] in 25 % 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 % HCA in Acetone: Olive oil 4:1 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

Positive control results:
The positive control group animals were treated with 25 % (w/v) HCA solution (dissolved in AOO) concurrent to the test item groups. No mortality, cutaneous reactions or signs of toxicity were observed in the positive control group.
Significant lymphoproliferative response (SI ≥ 3) was noted for HCA (SI = 8.1). The results of the positive control item demonstrated appropriate performance of the test in accordance with the relevant guidelines and confirmed validity of the assay.

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. Based on the results EC3 value (dose calculated to induce a stimulation index of 3) of the test item was not calculated.Dose-response relationship was evaluated by linear regression. 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.)

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

17750

17710.5

2213.8

1.0

Positive control: 25% HCA in AOO 

144132

144092.5

18011.6

8.1

 IMIDAZOLE ETHANOL

25% in DMF

16576

16536.5

2067.1

2.3

 IMIDAZOLE ETHANOL

10% in DMF

11831

11791.5

1473.9

1.6

 IMIDAZOLE ETHANOL

5% in DMF

14333

14293.5

1786.7

2.0

 IMIDAZOLE ETHANOL

2.5% in DMF

8751

8711.5

1088.9

1.2

 Vehicle control

for the test item: DMF

7226

7186.5

898.3

1.0

 

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Remarks:
Migrated information
Conclusions:
Under the conditions of the present Local Lymph Node Assay, “IMIDAZOLE ETHANOL” (CAS No. 24155-42-8) tested at the maximum attainable concentration of 25 % (w/v, based on solubility) and at concentrations of 10 %, 5 % and 2.5 % as formulations in an appropriate vehicle (DMF) was shown to have no sensitization potential.