1-Ethoxy-2,3-difluoro-4-[trans-4-(trans-4-propylcyclohexyl)-cyclohexyl]-benzene

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Jul 02 - Sep 26, 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

4-(4-ethoxy-2,3-difluorophenyl)-4'-propyl-1,1'-bi(cyclohexane)

EC Number:

602-941-8

Cas Number:

123560-48-5

Molecular formula:

C23H34F2O

IUPAC Name:

4-(4-ethoxy-2,3-difluorophenyl)-4'-propyl-1,1'-bi(cyclohexane)

Test material form:

solid

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Netherlands B.V. Postbus 6174 NL - 5960 AD Horst
- Age at study initiation: 7 - 8 weeks
- Weight at study initiation: 19.4 +/- 0.8 g
- Housing: single
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 30-85
- Photoperiod (hrs dark / hrs light): 12/12
IN-LIFE DATES: From: day 1 To: day 5

Study design: in vivo (LLNA)

Vehicle:

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

Concentration:

pre test for signs of irritation: 1, 2.5, 5, and 10 % (v/v)
main test: 2.5, 5, and 10 % (v/v)

No. of animals per dose:

pre test: 2
main test: 5 (f) per group (3 test groups, 1 control group) total 20 (f)

Details on study design:

RANGE FINDING TESTS:
To determine the highest non-irritant test concentration or the highest technically applicable concentration, a pretest was performed in two mice on three consecutive days. The data of a solubility experiment showed that the highest test item concentration, which could be technically used was a 10 % solution. At concentrations of 1 and 2.5 % the treated mouse did not show any signs of irritation. At concentrations of 5 and 10 % the treated mouse showed reddening of the ear skin only after the third application of the test item. The test item in the main study was, therefore, assayed at 2.5, 5, and 10 %.


MAIN STUDYANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: OECD 429
- Criteria used to consider a positive response: 3-fold greater response at one concentration in SI than control animals

TREATMENT PREPARATION AND ADMINISTRATION:
a) Topical application of 25 µL test item preparation (test group) or vehicle (control group)
b) five days after topical application: iv application of 3H-methyl thymidin
c) prior each treatment (a and b) ear thickness measurement
d) five hours after treatment (b) necropsy and analysis of the 3H thymidin incorporation in draining lymph nodes
e) method: pooled per animal

Positive control substance(s):

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

Statistics:

Dunnets Test

Results and discussion

Positive control results:

Conc. SI
5 %: 2.43
10 % 4.07
25 % 4.88

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

1.6

Test group / Remarks:

Test Group 2.5 %

Results 2

Key result

Parameter:

SI

Value:

2.02

Test group / Remarks:

Test Group: 5 %

Results 3

Key result

Parameter:

SI

Value:

1.12

Test group / Remarks:

Test Group: 10 %

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
DPM per lymph node
Control: 793.6
2.5 %: 1267.1
5 %: 1599.8
10 %: 889.4

DETAILS ON STIMULATION INDEX CALCULATION
ratio of 3HTdR incorporated into lymph node cells of test lymph nodes relative to that recorded for control lymph nodes

EC3 CALCULATION
The EC3 value could not be calculated since all SIs were below 3.

CLINICAL OBSERVATIONS
No deaths occurred during the study period.

BODY WEIGHTS
The body weight of the animals, recorded prior to the first application and prior to treatment with 3HTdR, was within the range commonly recorded for animals of this strain and age.

SIGNS OF TOXICITY
No symptoms of local toxicity at the ears of the animals and no systemic findings were observed during the study period.
The measured ear thickness of all animals treated was recorded prior to the 1stapplication and prior to treatment with 3HTdR. A relevant increase in ear thickness gain was not observed after treatment with the test item.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The test item was concluded to be no skin sensitiser.

Executive summary:

In the study the test item was dissolved in acetone:olive oil (4+1) was assessed for its possible contact allergenic potential. For this purpose a local lymph node assay was performed using test item concentrations of 2.5, 5, and 10 %. The GLP study was performed according to OECD 429. The animals did not show any clinical signs during the course of the study and no cases of mortality were observed.  A relevant increase in ear thickness gain could not be observed after treatment with the test item. In this study Stimulation Indices (S.I.) of 1.60, 2.02, and 1.12 were determined with the test item at concentrations of 2.5, 5, and 10 % in acetone:olive oil (4 +1), respectively. Since the S.I. was not increased above 3 at any test group, and EC3 could not be determined. From the data it can be concluded that the test item is not a skin sensitiser under the conditions of this assay.