2,2-dimethyl-1,3-propanediyl didecanoate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13 June - 21 July 2006

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

adopted in 1992

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Version / remarks:

adopted in 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Centre international de toxicologie (CIT), Miserey, France

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Test was done before LLNA as first-choice method for in-vivo testing was set into force.

Test material

Test material information

Constituent 1

Reference substance name:

2,2-dimethyl-1,3-propanediyl didecanoate

EC Number:

248-688-3

EC Name:

2,2-dimethyl-1,3-propanediyl didecanoate

Cas Number:

27841-06-1

Molecular formula:

C25H48O4

IUPAC Name:

3-(decanoyloxy)-2,2-dimethylpropyl decanoate

In vivo test system

Test animals

Species:

guinea pig

Strain:

Hartley

Remarks:

Crl: (HA) BR

Sex:

male/female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories France, L'Arbresle, France
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 1 - 2 months
- Weight at study initiation: 346 - 388 (males), 347 - 390 (females)
- Housing: individual, in polycarbonate cages with stainless steel lid, bedding: autoclaved sawdust
- Diet: 106 pelleted diet (SAFE, Villemoisson, Epinay-sur-Orge, France), ad libitum
- Water: filtered water (FG Millipore membrane: 0.22 micron), ad libitum (analyses were performed)
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 2
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

corn oil

Concentration / amount:

25%
0.1 mL/ injection site

Day(s)/duration:

single injection

Adequacy of induction:

other: highest concentration, which caused only moderate irritation in preliminary experiment

Induction 2

Route:

epicutaneous, occlusive

Vehicle:

unchanged (no vehicle)

Concentration / amount:

100%
closed patch, fully-loaded with test substance

Day(s)/duration:

48 h

Adequacy of induction:

other: pretreatment with 0.5 mL of 10% sodium lauryl sulfate (in vaseline); local irritation was observed in animals of control and treatment group

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

unchanged (no vehicle)

Concentration / amount:

100%
on filter paper of a chamber (Finn Chamber®), fully-loaded with test substance

Day(s)/duration:

24 h

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

Preliminary study: 4 animals (2 per sex)

Main study:
Test group: 20 (10 per sex)
Control group: 10 (5 per sex)

Details on study design:

RANGE FINDING TESTS: Dermal effects were evaluated by treating 2 animals (m/f) with 5%, 10% or 25% of the test substance for intradermal induction and 50% and 100% for epicutaneous induction and retreatment of the test substance (challenge).

Intradermal: Minimal irritation was observed at all tested concentrations of the test substance, which was augmented when the test substance was injected in combination with FCA (Freund's Complete Adjuvant), persisting until the last reading time point (Day 6).
Dermal: No signs of irritation was observed at none of the concentrations.
After treatment with 50% of the test substance in the challenge phase, the female guinea pig showed a discrete or patchy erythema at the 24-hour reading time point, which cleared until 48 h after patch removal.
Based on the results of the preliminary study the minimal irritating concentrations, 25% dilution of the test substance in corn oil was used for intradermal induction and 100% of the test substance was used for the epidermal induction exposure in the main study. 100% of the test substance was selected as maximally non-irritating concentration for the challenge phase.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 (intradermal and epicutaneous)
- Exposure period: single application (intradermal induction on Day 1) and 48 hours (epidermal induction on Day 8)
- Test group: 20 animals (f/m)
Intradermal (double injections):
Injection 1: 0.1 mL 1:1 mixture (v/v) FCA and 0.9% NaCl (FCA mix)
Injection 2: 0.1 mL test substance (25% in corn oil)
Injection 3: 0.1 mL test substance (25% in corn oil) in a 1:1 mixture (v/v) with FCA mix
Epicutaneous: sodium lauryl sulfate (10% in vaseline) (Day 7), test substance (100%) (Day 8)

- Vehicle control group: 10 animals (f/m)
Intradermal (double injections):
Injection 1: 0.1 mL 1:1 mixture (v/v) FCA and 0.9% NaCl (FCA mix)
Injection 2: 0.1 mL corn oil
Injection 3: 0.1 mL vehicle in a 1:1 mixture (v/v) with FCA mix
Epicutaneous: vehicle (corn oil)

- Site: symmetrically on both sides of the spine (interscapular region, right and left flank)
- Frequency of applications: once
- Concentrations: intradermal: 25%, epicutaneous: 100%
- Other: An examination of the injections was performed 24 and 48 h after treatment and on Day 6.

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: Day 22 (14 days after induction)
- Exposure period: 24 h
- Test group: test substance (right flank) and vehicle (corn oil) (left flank)
- Control group: test substance (right flank) and vehicle (corn oil) (left flank)
- Site: clipped and shaved posterior flanks
- Concentrations: 100% (undiluted)
- Evaluation (hrs after the end of the challenge exposure): 24 and 48 h

Challenge controls:

The control group is actually a challenge control.

Positive control substance(s):

yes

Remarks:

Mercaptobenzothiazole (tested separately)

Results and discussion

Positive control results:

Mercaptobenzothiazole was used as positive control in an independent experiment (March 2006) at a challenge concentration of 20% (in corn oil) (intradermal induction concentration: 1%; epicutaneous induction: 20%) 10/10 treated guinea pigs showed positive reactions.

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

intradermal induction: 25%; challenge: 100%

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Results 2

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

intradermal induction: 0%; challenge: 100%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

no indication of skin sensitisation

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

intradermal induction: 1%; challenge: 20%

No. with + reactions:

10

Total no. in group:

10

Clinical observations:

edema (8/10), dryness of skin (7/10), crusts (1/10), intense erythema (9/10), moderate and confluent erythema (1/10)

Remarks on result:

positive indication of skin sensitisation

Remarks:

(tested in an independent experiment)

Results 4

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

intradermal induction: 25%; challenge: 100%

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

In 3/20 animals a dryness of the skin was observed.

Remarks on result:

no indication of skin sensitisation

Results 5

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

intradermal induction:0%; challenge: 100%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

no indication of skin sensitisation

Results 6

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

intradermal induction: 1%; challenge: 20%

No. with + reactions:

10

Total no. in group:

10

Clinical observations:

edema (9/10), dryness of skin (5/10), intense erythema (3/10), moderate and confluent erythema (1/10), discrete or patchy erythema (1/10), reading was masked by a considerable dryness of the skin (5/10)

Remarks on result:

positive indication of skin sensitisation

Remarks:

(tested in an independent experiment)

Any other information on results incl. tables

Range finding test

Table 1: Results after intradermal injection

Animal No.	Concentration (w/w) [%]	Reaction scores after treatment
		24 h	48 h	6 d
1 (M)	25 (in corn oil / FCA mix (1:1))	I	I	I
	25 (in corn oil)	I	LI	LI
	10 (in corn oil / FCA mix (1:1))	I	I	I
	10 (in corn oil)	I	LI	LI
	5 (in corn oil / FCA mix (1:1))	I	I	I
	5 (in corn oil)	LI	LI	LI
2 (F)	25 (in corn oil / FCA mix (1:1))	I	I	I
	25 (in corn oil)	I	LI	LI
	10 (in corn oil / FCA mix (1:1))	I	I	I
	10 (in corn oil)	LI	LI	LI
	5 (in corn oil / FCA mix (1:1))	I	I	I
	5 (in corn oil)	LI	LI	LI

F = female

FCA mix = Freund’s Complete Adjuvant / 0.9% NaCl (50/50, v/v)

I = irritation

LI = slight irritation

M = male

Table 2: Results after epicutaneous application (erythema scores)

	Animal No.	Concentration (w/w) [%]	Reaction scores after removal of dressing
			24 h	48 h
Under conditions of challenge phase	1 (M)	100	0	0
		50	0	0
	2 (F)	100	0	0
		50	1	0
Under conditions of induction phase	3 (M)	100	0	0
	4 (F)	50	0	0

F = female

M = male

Main study

Results after intradermal injection and epicutaneous application of the test substance

No residual test substance was observed upon removal of the dressing on Day 10. A local irritation was recorded ind both, control and test group.

Challenge

Table 3: Skin response (erythema scores) after challenge

			Reaction scores after
		Animal No.	24 h		48 h
			LF	RF	LF	RF
Control group	Male	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
	Female	16	0	0	0	0
		17	0	0	0	0
		18	0	0	0	0
		19	0	0	0	0
		20	0	0	0	0
Test group	Male	6	0	0	0	0/S
		7	0	0	0	0
		8	0	0	0	0
		9	0	0	0	0/S
		10	0	0	0	0
		11	0	0	0	0/S
		12	0	0	0	0
		13	0	0	0	0
		14	0	0	0	0
		15	0	0	0	0
	Female	21	0	0	0	0
		22	0	0	0	0
		23	0	0	0	0
		24	0	0	0	0
		25	0	0	0	0
		26	0	0	0	0
		27	0	0	0	0
		28	0	0	0	0
		29	0	0	0	0
		30	0	0	0	0

LF = left flank (vehicle)

RF = right flank (100% test substance)

S = dryness

Mortality / viability/ systemic effects/ body weights

No mortalities or signs indicative for systemic or local toxicity were observed. Neither did the clinical observations reveal any abnormalities. The body weight gain of the animals was not affected adversely during the study. No necropsy was performed after the sacrifice of the animals.

Applicant's summary and conclusion

Interpretation of results:

other: CLP/ EU GHS criteria not met, no classification required according to Regulation (EC) No. 1272/2008