4-trans-(4-trans-Propylcyclohexyl)-cyclohexancarbonic acid

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2002-01-22 to 2002-02-15

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Report date:

2002

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Version / remarks:

Directive 96/54/EEC

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

07-1992

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Study was performed before development of the LLNA.

Test material

Test material information

Constituent 1

Reference substance name:

(1r,1's,4r,4'r)-4'-propyl-[1,1'-bi(cyclohexane)]-4-carboxylic acid

EC Number:

613-782-9

Cas Number:

65355-32-0

Molecular formula:

C16 H28 O2

IUPAC Name:

(1r,1's,4r,4'r)-4'-propyl-[1,1'-bi(cyclohexane)]-4-carboxylic acid

Test material form:

solid: particulate/powder

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Winkelmann GmbH, Borchen, Germany
- Age at study initiation: about 5 weeks
- Weight at study initiation: 316 - 375 g
- Housing: 2 animals per Makrolon cage
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 22
- Humidity (%): 47 - 62
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: Days 1 to 25

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

paraffin oil

Concentration / amount:

Group 1 (vehicle):
- cranial 0.1 mL Freud's complete adjuvant (FCA) + NaCl solution,
- medial 0.1 mL vehicle,
- caudal 0.1 mL FCA + NaCl solution

Group 2 (test item):
- cranial 0.1 mL FCA + NaCl solution,
- medial 0.1 mL test item (25 g/L in liquid paraffin),
- caudal 0.1 mL FCA with test item + NaCl solution (25 g test item/L preparation)

Day(s)/duration:

day 1, single application

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Induction 2

Route:

epicutaneous, occlusive

Vehicle:

polyethylene glycol

Concentration / amount:

Filter paper patch fully soaked with 1 mL of the test item preparation or vehicle.
Group 1 (vehicle): PEG 400
Group 2 (test item): Test item (200 g/L in PEG 400)

Day(s)/duration:

over 48 hours, one week after intradermal injection (day 8)

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

polyethylene glycol

Concentration / amount:

Fixing filter papers fully loaded with 0.5 mL of test item preparation or soaked with 0.5 mL PEG 400.
Group 1 (vehicle): PEG 400
Group 2 (test item): Test item (25 g/L in PEG 400)

Day(s)/duration:

over 24 hours, 2 weeks after topical induction (day 22)

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

Pretest: 5 females
Group 1 (negative controls): 5 females
Group 2 (test material group): 10 females

Details on study design:

RANGE FINDING TESTS: To determine the concentrations suitable for the main study, a pre-test with single intradermal or topical administrations of the vehicle and of the test item preparations were performed. Intradermal injections were given to one animal. Four further animals were used to determine the topical concentrations for the main study. Two of the guinea pigs were exposed for 48 hours without pre-treatment to determine the concentrations for the dermal induction. The other two animals were treated according to the control group including intradermal injections with FCA. One week after topical induction, the two animals of the pretest group were exposed for 24 hours to different test material concentrations to find out the challenge concentration. In the pre-test, the following concentrations were used:

1. Test item with liquid paraffin as vehicle:
- intradermal: 50, 25, 10, 5 and 1 g/L

2. Test item with polyethylene glycol 400 as vehicle:
- topical: 400, 200, 100 and 10 g/L
- topical with FCA: 150, 100, 50 and 10 g/L

MAIN STUDY
A. INDUCTION EXPOSURE
- No of exposures: 2 (One induction on day 1 (intradermal injection) and another one one week after intradermal injections (topical application))
- Exposure period: 3 weeks in total (including 2 weeks induction period)
- Test groups: 2 (vehicle and test item)
- Control group: liquid paraffin (intradermal injection) and polyethylene glycol (topical applications)
- Site: shoulder region, three treatments on each side of the spinal column
- Duration: single intradermal injection, after one week topical application for 48 h under occlusive conditions
- Concentrations: 25 g/L test item in 0.1 mL (intradermal injection); 200 g/L in 1 mL (topical application)

B. CHALLENGE EXPOSURE
- No of exposures: 1 (two weeks after topical induction)
- Exposure period: 24 hours with occlusive plastic tapes, then another 48 hours
- Test groups: 25 g/L test item in polyethylene glycol 400
- Control group: polyethylene glycol 400
- Site: shaven sides of the animals
- Concentrations: 25 g/L test item in 0.5 mL
- Evaluation (hr after challenge): 48 hours and 72 hours

Challenge controls:

as challenge control, 5 female guinea pigs were treated with polyethylene glycol 400

Positive control substance(s):

not required

Remarks:

The sensitivity of the test system had been demonstrated periodically. The control was carried out with alpha-Hexylcinnamaldehyde as a sensitizer

Results and discussion

Positive control results:

The sensitivity of the test system has to be demonstrated periodically. The control was carried out with α-Hexylcinnamaldehyde as a sensitiser.

In vivo (non-LLNA)

Results

Results 1

Reading:

2nd reading

Hours after challenge:

72

Group:

positive control

Remarks on result:

positive indication of skin sensitisation

Remarks:

The sensitivity of the test system has to be demonstrated periodically. The control was carried out with α-Hexylcinnamaldehyde as a sensitiser.

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

72

Group:

test chemical

Dose level:

25 g/L

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

no

Remarks on result:

no indication of skin sensitisation

Results 3

Key result

Reading:

2nd reading

Hours after challenge:

72

Group:

negative control

Dose level:

0 g/L

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

no

Remarks on result:

no indication of skin sensitisation

Results 4

Reading:

1st reading

Hours after challenge:

48

Group:

positive control

Remarks on result:

positive indication of skin sensitisation

Remarks:

The sensitivity of the test system has to be demonstrated periodically. The control was carried out with α-Hexylcinnamaldehyde as a sensitiser.

Results 5

Key result

Reading:

1st reading

Hours after challenge:

48

Group:

test chemical

Dose level:

25 g/L

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

no

Remarks on result:

no indication of skin sensitisation

Results 6

Key result

Reading:

1st reading

Hours after challenge:

48

Group:

negative control

Dose level:

0 g/L

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

no

Remarks on result:

no indication of skin sensitisation

Any other information on results incl. tables

Table 1:Findings after challenge

Group	Induction with	Challenge with	Positive animals (h after challenge)		Positive animals overall
			48 h	72 h
1	Vehicles*	PEG 400	0/5	0/5	0/5
		Test item #	0/5	0/5	0/5
2	Test item	Test item	0/10	0/10	0/10
		PEG 400	0/10	0/10	0/10

Vehicles           * = liquid paraffin and PEG 400

# = testing for primary irritation

 

Table 2. Body weight

	Mean body weight in g on experimental day					Difference day 1 to 25
Group	1	8	15	22	25
1 (vehicle)	332	395	441	481	502	+170
2 (test material)	353	419	471	529	543	+190

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In an in vivo skin sensitisation study according to OECD guideline 406, the test item showed no positive responders and has not to be classified as a skin sensitizer.

Executive summary:

The test item was evaluated for skin sensitizing properties in the guinea pig maximization test according to Magnusson and Kligman (1969). 5 female guinea pigs in the negative control group (group 1) treated with the vehicles (liquid paraffin and polyethylene glycol 400) and 10 females in the test item group (group 2) were investigated. Induction included intradermal injection of the test item preparation in liquid paraffin (25 g/L with and without Freud's complete adjuvant) on experimental day 1, and topical apllication of the test item preparation in polyethylene glycon 400 (200 g/L) on day 8 of the experimental part. Challenge by topical application of the test item preparation on polyethylene gylcol 400 (25 g/L) was performed two weeks after topical induction. After challenge, no positive reactions in the test item treated skin sites at both readings (24 and 72 h after start of challenge) were seen.