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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-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 guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
May 2008
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
adopted 17th July 1992
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
A valid GPMT conducted according to guideline is available, which is reliable without restrictions and adequate for classification and labelling purposes. Potency estimation is not mandatory when existing guideline and GLP conforming data are available, which were conducted before the new annex of the REACH Regulation entered into force. Moreover, no indication for skin sensitisation was observed in this study, thus, no dose response information is needed. For this reason and for reasons of animal welfare no additional LLNA was conducted.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material: MDIPA-Esterquat C16-18 and C18 unsatd.
- Physical state: solid
- Analytical purity: 100%

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River France, L’arbresle Cedex, France
- Age at study initiation: approx. 8 weeks
- Housing: Group housing of maximally 5 animals per labeled Noryl cage (Tecniplast; 74 cm x 54 cm x 25 cm height) containing sterilized sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France) and shelters (CS3B02A Play tunnels (90 mm x 5 mm x 125 mm), Datesand, Manchester, UK) as cage enrichment
- Diet (e.g. ad libitum): Complete maintenance diet for guinea pigs (SSNIFF® Spezialdiäten GmbH, Soest, Germany) ad libitum; hay (TecniLab-BMI BV, Someren, The Netherlands) was provided at least twice a week
- Water (e.g. ad libitum): tap weater, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-24
- Humidity (%): 40-70
- Air changes (per hr ):approx. 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
water
Concentration / amount:
Preliminary irritation study
Intradermal: 10, 5, 2, 1, 0.5, 0.2, 0.1, 0.05%
Epidermal: 50, 40, 30, 20, 10, 5, 2, 1%

Main study
Intradermal induction: 0.2%
Epidermal induction: 10%
Challenge: 1%
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
water
Concentration / amount:
Preliminary irritation study
Intradermal: 10, 5, 2, 1, 0.5, 0.2, 0.1, 0.05%
Epidermal: 50, 40, 30, 20, 10, 5, 2, 1%

Main study
Intradermal induction: 0.2%
Epidermal induction: 10%
Challenge: 1%
No. of animals per dose:
Experimental group: 10 females
Control group: 5 females
Details on study design:
RANGE FINDING TESTS:
A preliminary irritation study was conducted in order to select test substance concentrations to be used in the main Study. The selection of concentrations was based on the following criteria:
-The concentrations are well-tolerated systemically by the animals.
-For the induction exposures: the highest possible concentration that produced mild to moderate irritation (grades 2 - 3).
-For challenge exposure: the maximum non-irritant concentration.
Intradermal: 10, 5, 2, 1, 0.5, 0.2, 0.1, 0.05%
Epidermal: 50, 40, 30, 20, 10, 5, 2, 1%

Based on the results, the test substance concentrations selected for the main study were a 0.2% concentration for the intradermal induction and a 10% concentration for the epidermal induction exposure. A 1% test substance concentration was selected for the challenge phase.

MAIN STUDY
A. INDUCTION EXPOSURE
intradermal
- Test groups:
A) A 1:1 w/w mixture of Freunds Complete Adjuvant with water for injection
B) The test substance at a 0.2% concentration
C) A 1:1 w/w mixture of the test substance, at twice the concentration used in (B) and Freunds Complete Adjuvant
- Control group: animals were treated as described for the experimental animals except that, instead of the test substance, vehicle alone was administered
- Site: scapular region
- Frequency of applications: 1x

epidermal
- No. of exposures: 1
- Exposure period: 48 h
- Test groups: 0.5 mL of a 10% test substance concentration
- Control group: 0.5 mL vehicle
- Site: scapular area
- Frequency of applications: 1x


B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 24, 25
- Exposure period: 24 h
- Test groups: 0.1 mL 1% test substance, 0.1 mL vehicle
- Control group: 0.1 mL 1% test substance, 0.1 mL vehicle
- Site: flank
- Evaluation (hr after challenge): 24, 48 h
Positive control substance(s):
yes
Remarks:
Alpha- Hexylcinnamaldehyde

Results and discussion

Positive control results:
A reliability check with the positive control substance Alpha- Hexylcinnamaldehyde is carried out at regular intervals (last check Jan/Feb 2013) to check the sensitivity of the test system.
Concentrations selected for this study were:
Intradermal induction: A 20% solution in water (w/w).
Epidermal induction: undiluted.
Challenge: a 20% solution in water (w/w).
The positive control did induce skin sensitisation.

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
1%
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 1%. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
1%
No. with + reactions:
1
Total no. in group:
10
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 1%. No with. + reactions: 1.0. Total no. in groups: 10.0.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
1%
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 1%. No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
1%
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 1%. No with. + reactions: 0.0. Total no. in groups: 5.0.

Any other information on results incl. tables

Preliminary irritation study

SKIN REACTIONS AFTER INTRADERMAL INJECTION

Animal

Conc %

24 hours after injection

48 hours after injection

number

 

Erythema

Necrosis

Erythema

Necrosis

 

 

(grade)

(mm)

(grade)

(mm)

48

   10

 

5

 

6

 

     5

 

4

 

5

49

     2

 

3

 

4

 

     1

 

2

 

3

45

0.5

3

 

 

1

 

0.2

2

 

1

 

50

0.1

2

 

1

 

 

0.05

1

 

1

 

 

SKIN REACTIONS AFTER EPIDERMAL EXPOSURE

Animal

Conc. %

24 hours after exposure

48 hours after exposure

number

 

Erythema

Oedema

Erythema

Oedema

 

 

(grade)

(grade)

(grade)

(grade)

46

10

0

0

0

0

 

5

0

0

0

0

47

10

0

0

0

0

 

5

0

0

0

0

48

2

0

0

0

0

 

1

0

0

0

0

49

2

0

0

0

0

 

1

0

0

0

0

45

50

N

1

N

2

 

40

N

1

N

2

50

30

N

1

N

2

 

20

N

0

N

1

Main Study

Induction readings

Animal number

Intradermal injection (Day 3)

Epidermal exposure (Day 10)

 

 

A

B

D

D

 

Control group

E

N

E

N

E

N

Erythema

Oedema

 

   31

3

 

0

 

3

 

0

0

   32

3

 

1

 

2

 

0

0

   33

3

 

1

 

3

 

0

0

   34

3

 

0

 

3

 

0

0

   35

3

 

0

 

3

 

0

0

Experimental group

E

N

E

N

E

N

 

 

   36

3

 

2

 

3

 

3

0

   37

3

 

3

 

3

 

3

0

   38

3

 

2

 

2

 

3

0

   39

3

 

2

 

3

 

3

0

   40

3

 

3

 

3

 

3

0

   41

3

 

3

 

3

 

3

0

   42

3

 

3

 

3

 

3

0

   43

3

 

3

 

3

 

3

0

   44

3

 

3

 

3

 

3

0

   45

3

 

3

 

3

 

3

0

 

A. 1:1 Mixture of Freund’s Complete Adjuvant and water for injection.

B. A 0.2% test substance concentration (Experimental); vehicle (Control).

C. 1:1 Mixture of Freund’s Complete Adjuvant and a 0.4% concentration (Experimental) or vehicle (Control).

D. A 10% test substance concentration (Experimental); vehicle (Control).

 

Skin effects intradermal injections:

E. Erythema (grade)

N. Signs of necrosis (mm in diameter)


Challenge readings

Animal number

DAY 24

DAY 25

 

1%#

Vehicle*

1%#

Vehicle*

Control group

     1

    0

    0

    0

    0

     2

    0

    0

    0

    0

     3

    0

    0

    0

    0

     4

    0

    0

    0

    0

     5

    0

    0

    0

    0

Experimental group

   36

    0

    0

    0

    0

   37

    0

    0

    0

    0

   38

    0

    0

    0

    0

   39

    0

    0

    0

    0

   40

    0

    0

    0

    0

   41

    0

    0

    0

    0

   42

    0

    0

    1p

    0

   43

    0

    0

    0

    0

   44

    0

    0

    0p

    0

   51

    0

    0

    0

    0

 

#. Test substance concentration.

 *. Water (Elix)

p. Scaliness


Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
In this Guinea Pig Maximisation Test, MDIPA-Esterquat C16-18 and C18 unsatd. was not sensitising.
Executive summary:

In a dermal sensitisation study according to OECD Guideline 406 (adopted 17th July 1992) and EU method B.6 (May 2008) with MDIPA-Esterquat C16-18 and C18 unsatd. (100% a.i.) in water, young adult female Dunkin-Hartley guinea pigs (10 in test group, 5 in control group) were tested using the method of Magnusson and Kligman. Alpha-Hexylcinnamaldehyde was used as positive control.

Based on the results of a preliminary study, the test substance concentrations selected for the main study were a 0.2% concentration for the intradermal induction and a 10% concentration for the epidermal induction exposure. A 1% test substance concentration was selected for the challenge phase.

After epidermal induction all animals of the experimental group showed signs of irritation. Following a challenge exposure to a 1% test substance concentration, one experimental animal showed discrete or patchy erythema, with scaliness, and one experimental animal showed scaliness only, both at 48 hours after exposure. No skin reactions were evident in the control animals and all other experimental animals. The sensitisation rate was 10%.

According to CLP, EU GHS (Regulation (EC) No 1272/2008), a response of at least 30% of the test animals of an adjuvant type guinea pig test method for skin sensitisation is considered as positive.

MDIPA-Esterquat C16-18 and C18 unsatd. is not a dermal sensitiser in this study.