Dodecanoic acid, 1,1'-[1,6-hexanediylbis[nitrilo(2,2-dimethyl-1-propanyl-3-ylidene)]] ester

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2005-02-16 to 2005-06-16

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Version / remarks:

1996

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

1992

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Version / remarks:

1996

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

The Guinea Pig Maximazitation Test (2005) met the previous requirements before the entry into force of REACH. The GPMT is suitable and reliable to cover this endpoint. For this reason and for animal welfare reasons, no further in vivo study (LLNA test) needs to be performed.

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

479-930-8

EC Name:

-

Cas Number:

613222-52-9

Molecular formula:

C40H76N2O4

IUPAC Name:

(3E)-3-({6-[(E)-[3-(dodecanoyloxy)-2,2-dimethylpropylidene]amino]hexyl}imino)-2,2-dimethylpropyl dodecanoate

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: LAB-ÁLL Bt. Budapest, 1174 Hunyadi u. 7.
- Age at study initiation: 29 days;
- Weight at study initiation: mean 336.5 grs;
- Housing: macrolon cages, size III., with 3 or 2 animals/cage (42 x 42 x 19 cm);
- Diet: PURISTAR standard diet for rabbits produced by AGRIBRANDS Europe Hungary, 7400 Kaposvár, Dombóvári út, ad libitum.
- Water: tap water, as for human consumption, ad libitum supplemented with 50 mg/100 ml Ascorbic acid.
- Acclimation period: 29 days

ENVIRONMENTAL CONDITIONS
- Temperature: 20 ± 3 °C
- Humidity: 30 - 70 % R.H.
- Air changes: 8 - 12 air exchange/hour by central air-condition system
- Photoperiod: 12 hours daily from 6 a.m. to 6 p.m. (artificial light)

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

other: Oleum helianthi

Concentration / amount:

5%

Day(s)/duration:

Day 1

Adequacy of induction:

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

Induction 2

Route:

epicutaneous, occlusive

Vehicle:

other: Oleum helianthi

Concentration / amount:

100%

Day(s)/duration:

Day 8/ 48 hours

Adequacy of induction:

non-irritant substance, but skin pre-treated with 10% SDS

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

other: Oleum helianthi

Concentration / amount:

100%

Day(s)/duration:

Day 22/ 24 hours

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

10 animals in the treatment group;
5 control animals;

Details on study design:

RANGE FINDING TESTS:
A. INTRADERMAL INJECTION
- Concentrations: 0.01, 0.1, 1 and 5% (w/v) in oleum helianthi
- Site: right and left flanks (each 2 concentrations)

B. TOPICAL APPLICATION
- Concentrations: 25, 50, 75% (w/v) in oleum helianthi and undiluted
- occlusive bandage
- two animals were each exposed to two different compounds

Results:
Intra-dermal treatment with test item caused local irritation at concentrations of 1 and 5% up to 48 hours and 24 hours, respectively. At a concentration of 5% well defined erythema occured in both animals one hour after patch removal (score 2). At concentration of 1% well defined and very slight erythema occured one hour after patch removal (score 1). At concentrations of 0.01 and 0.1% no primary irritation was observable (score 0). Topical application of 0.5 mL test formulation at concentrations of 25, 50, 75% and in undiluted state produced no reaction on the skin of guinea pigs (score 0) Therefore, the test item was used at a concentration of 5% for intra-dermal treatment and used in undiluted state for dermal induction treatment and challenge application. The test area was exposed to 0.5 mL of 10% SDS in vaseline 24 hours prior to topical induction applications, in order to create local irritation.

MAIN STUDY
A. 1. INDUCTION EXPOSURE (Injection); Day 1
- No. of exposures: series of three injection to each side (0.1 mL)
- Test groups: 5% test item in oleum helianthi; Freund’s complete adjuvant (FCA) mixed with physiological saline (1:1); 5% test item mixed with physiological saline solution and homogenized in Freund’s complete adjuvant
- Control group: Oleum helianthi; mix of Freund’s complete adjuvant and physiological saline (1:1) (v/v); 50 v/v % oleum helianthi in a 1:1 mixture (v/v) of Freund’s complete adjuvant and physiological saline
- Site: scapular region
- Frequency of applications: once

A.2. INDUCTION EXPOSURE (Epicutaneous); Day 8
- Exposure period: 48 hours
- Test groups: undiluted test item in oleum helianthi
- Control group: oleum helianthi only
- Frequency of applications: once

B. CHALLENGE EXPOSURE (Epicutaneous)
- Day(s) of challenge: Day 22
- Exposure period: 24 hours
- Test groups and control group: undiluted test item (left flank) and vehicle (right flank)
- Site: left shaved flank (undiluted test item), right shaved flank (vehicle only)
- Evaluation (hr after challenge): 24 and 48

Challenge controls:

Concurrent to treatment animals, 5 control guinea pigs were exposed to the vehicle only, during the sensitisation phase, and treated with the test item (in undiluted state) in challenge exposures.

Positive control substance(s):

yes

Remarks:

potassium dichromate

Results and discussion

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

100%

No. with + reactions:

4

Total no. in group:

10

Clinical observations:

moderate to intense erythema and swelling

Remarks on result:

positive indication of skin sensitisation

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

undiluted

No. with + reactions:

4

Total no. in group:

10

Clinical observations:

moderate to intense erythema and swelling

Remarks on result:

positive indication of skin sensitisation

Results 3

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

100%

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

no indication of skin sensitisation

Results 4

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

100%

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

no indication of skin sensitisation

Results 5

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

0.3%

No. with + reactions:

5

Total no. in group:

10

Remarks on result:

positive indication of skin sensitisation

Results 6

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

0.3%

No. with + reactions:

5

Total no. in group:

10

Remarks on result:

positive indication of skin sensitisation

Any other information on results incl. tables

Dermal response scores for Guinea pigs challenged with the test item:

ANIMAL (test group)	SCORES OF DERMAL REACTION		ANIMAL (control group)	SCORES OF DERMAL REACTION
NUMBER	24 hours	48 hours	NUMBER	24 hours	48 hours
	after the patch removal			after the patch removal
263	0	0	261	0	0
266	0	0	262	0	0
268	3	2	264	0	0
269	0	0	265	0	0
270	0	0	267	0	0
271	3	3	-	-	-
272	2	2	-	-	-
273	0	0	-	-	-
274	2	2	-	-	-
275	0	0	-	-	-
number of positive/ number of tested	4/10	4/10	number of positive/ number of tested	0/5	0/5

 

 

Scoring of Skin Sensitisation

 0      = no visible change

1      = discrete or patchy erythema

2      = moderate and confluent erythema

3      = intense erythema and swelling

 

Applicant's summary and conclusion

Interpretation of results:

Category 1B (indication of skin sensitising potential) based on GHS criteria

Conclusions:

The test item was classified as a skin sensitiser, based on the guinea pig maximisation test and according to current EU-regulations.

Executive summary:

SIKA Hardener LH was tested for skin sensitising properties in a guinea pig maximisation test according to EU method B.6 and OECD guideline 406. A guinea pig maximisation test (GPMT) skin sensitisation study was performed according to the Magnusson and -Kligman method, using Freund's complete adjuvant technique, to evaluate the sensitisation potential of test item SIKA Hardener LH.

A total of 10 test animals were subjected to sensitisation procedures in a two-stage operation, i.e. an intra-dermal treatment and a topical application. The test item was administered at a concentration of 5 % for intra dermal injections and in undiluted state for dermal sensitisation treatments. The test area was painted with 0.5 ml of 10 % sodium dodecyl sulphate in vaseline 24 h prior to the topical induction application in order to create a local irritation. Two weeks following the last induction exposure a challenge dose (in undiluted state) was administered. Challenge was performed by dermal application of the test item.

Concurrent to treatment animals, 5 control guinea pigs were exposed to the vehicle only, during the sensitisation phase, and treated with the test item (in undiluted state) in challenge exposures.

A response, indicating sensitising effects, was observed in 40 % of the treatment group animals after challenge application of the test item. Challenge with the test item elicited intense, confluent erythema and swelling (scores 2-3) in guinea pigs previously sensitized with test item. No response was observed in concurrently challenged control group animals. In summary, SIKA Hardener LH was considered sensitising to skin.