N,N-dimethyldecylamine N-oxide

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 november 2011-10 February 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

yes

Remarks:

In the main study the skin was coated with sodium lauryl sulfate (SLS) on the day before stage 2 induction in order to ensure the induction of local irritation.

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

yes

Remarks:

In the main study the skin was coated with sodium lauryl sulfate (SLS) on the day before stage 2 induction in order to ensure the induction of local irritation.

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

The decision to perform this study in preference to the LLNA was taken based on the surface active properties of the substance. The potential for surface active substances to demonstrate sensitizing properties has been shown to be more accurately predicted using the M&K procedure than the LLNA. Some surfactants have been shown to demonstrate false positive responses in the LLNA, for example sodium lauryl sulfate.

Test material

Test material information

Constituent 1

Reference substance name:

N,N-dimethyldecylamine N-oxide

EC Number:

220-020-5

EC Name:

N,N-dimethyldecylamine N-oxide

Cas Number:

2605-79-0

Molecular formula:

C12H27NO

IUPAC Name:

decyl(dimethyl)amine oxide

Test material form:

other: aqueous solution

Details on test material:

- Name of test material (as cited in study report): N,N-dimethyldecylamine-N-oxide (solution)
- Substance type: colourless to yellowish liquid
- Analytical purity: 40.5% active
- Impurities (identity and concentrations): peroxide: 0.01%, free amine: < 0.5%
- Purity test date: 2011-11-14
- Lot/batch No.: 1108042301
- Expiration date of the lot/batch:2 years in original closed packaging
- Storage condition of test material: 10 - 25°C
- Other: pH = 7.59

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services Germany GmbH, Stolzenseeweg 32 – 36, 88353 Kißlegg, Germany
- Age at study initiation: 42 days
- Weight at study initiation: 315-420 g (excluding positive control group); positive control group 332-364 g
- Housing: The animals were kept in pairs in MAKROLON cages (MZK 80/25)
- Diet: Commercial diet, ssniff® Ms-H V2233 (ssniff Spezialdiäten GmbH, 59494 Soest, Germany) served as food. This food was offered ad libitum.
- Water: Tap water (in drinking bottles) was offered ad libitum.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 15
- Air changes (per hr): no information
- Photoperiod (hrs dark / hrs light): 12h/12h

IN-LIFE DATES: From: To: 28-11-2011 - 10-02-2012

Study design: in vivo (non-LLNA)

Induction

Route:

intradermal and epicutaneous

Vehicle:

water

Concentration / amount:

Preliminary Study:
Induction Phase: Intracutaneous - 0.1 mL of 0.004, 0.04, 0.4, 2 or 4% C10 AO; Topical - 2 mL of 0.4, 2, 4, 10, 20, 30 or 40 % C10 AO (non-depilated and depilated skin) and in addition 2 mL of 0.004, 0.02, 0.04, 0.2% C10 AO (depilated skin)
Main Study:
Induction Phase: Stage 1 (intracutaneous) - 0,1 mL of 0.2% C10 AO; Stage 2 (topical) - 2 mL of 10% C10 AO

Adequacy of induction:

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

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

2 mL of 0.04% C10 AO

Adequacy of challenge:

not specified

No. of animals per dose:

Preliminary Study: 10 animals for topical and 2 animals for intracutaneous
Main study: 20 animals for C10 AO; 10 animals for vehicle control; Historical data used for positive controls

Details on study design:

RANGE FINDING TESTS: see below

MAIN STUDY
A. INDUCTION EXPOSURE
STAGE 1 - INTRACUTANEOUS
- No. of exposures:1
- Exposure period: Day 0
- Test groups: (1) FCA (diluted 1:1 with 0.9% NaCl); (2) 0.2% C10 AO; (3) 0.2% C10 AO in a 1:1 mixture with FCA/physiological saline
- Control group: -
- Site: Shoulder
- Frequency of applications: single application
- Duration: -
- Concentrations: see above

STAGE 2 - TOPICAL
- No. of exposures:1
- Exposure period: Day 7
- Test groups: 10% C10 AO
- Control group: -
- Site: Shoulder
- Frequency of applications: single application
- Duration: -
- Concentrations: see above

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: Day 21
- Exposure period: 24 hrs
- Test groups: 10% C10 AO
- Control group: tap water (vehicle control)
- Site: L flank (test substance) R flank (control)
- Concentrations:
- Evaluation: 24 and 48 hrs after challenge

OTHER: On Day 6 the exposed skin was coated with 0.5 mL sodium lauryl sulfate (10% in vaseline) in order to ensure the induction of local irritation.

Challenge controls:

Treated with 0.04% C10 AO solution

Positive control substance(s):

yes

Remarks:

alpha-hexyl cinnamic aldehyde

Results and discussion

Positive control results:

Animals of the same strain treated with alpha-hexyl cinnamic aldehyde in sesame oil exhibited a sensitising reaction in all animals in form of a discrete or patchy erythema (grade 1).

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

0.04 %

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

no animals with positive responses

Results 2

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

0.04 %

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

no animals with positive responses

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

100%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: vehicle control

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

100%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: vehicle control

Results 5

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

0.01%

No. with + reactions:

20

Total no. in group:

20

Remarks on result:

other: historical background group from October/November 2011

Results 6

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

0.01%

No. with + reactions:

20

Total no. in group:

20

Remarks on result:

other: historical background group from October/November 2011

Any other information on results incl. tables

Preliminary Study:

Six concentrations of C10 AO were tested by intracutaneous injection: 0.004, 0.04, 0.2, 0.4, 2 or 4% solutions in tap water: A concentration of 0.004% revealed a discrete or patchy erythema, concentrations of 0.04% or 0.2% revealed a moderate and confluent erythema 24 to 72 hours after administration, respectively. Concentrations of 0.4%, 2% or 4% revealed an intense erythema and swelling with necrosis 24 to 72 hours after administration.

Several concentrations of C10 AO were tested by topical application: 0.4, 2, 4, 10, 20, 30 and 40% solutions to the non-depilated and depilated skin and, in addition, 0.004, 0.02, 0.04 and 0.2% to the depilated skin.

Non-depilated skin

Concentrations of 0.4% to 10% revealed a moderate and confluent erythema 48 hours and a discrete or patchy erythema 72 hours after start of exposure. The animals that received the 20% + 30% or 40% concentrations (equivalent to 3,300 and 2,640 mg AO/kg respectively) died prematurely. The cause of death could not be determined in this study.

Depilated skin

No skin reactions were observed up to the concentration of 0.04%. The concentration of 0.2% revealed a discrete or patchy erythema, concentrations of 0.4% or 2% revealed a moderate and confluent erythema 24 to 72 hours after start of exposure, respectively. Concentrations of 4% or 10% revealed a moderate and confluent erythema with brownish-green spots on the skin 24 to 72 hours after start of exposure. The animals receiving the 20% + 30% or 40% concentration (equivalent to 3,300 and 2,640 mg AO/kg respectively) died prematurely.

As mentioned, it was not possible to determine the cause of death of the animals in this study. In a subsequent study performed according to OECD TG 402 (see section 7.2.3) the acute dermal toxicity to rats was determined to be >2000 mg AO/kg bw, with no deaths or signs of toxicity observed in the study.

Based on the results of the preliminary study, it was decided to use a 0.2% concentration in tap water for the 1st (intracutaneous) induction stage, a 10% concentration in tap water for the 2nd (topical) induction stage and a 0.04% solution in tap water for the challenge.

Main Study:

A 0.2% solution of C10 AO chosen for the first (intracutaneous) induction stage revealed a discrete or patchy erythema in all 20 animals 24 and 48 hours after administration. However, in the main study the skin was coated with sodium lauryl sulfate on the day before the second (topical) induction using 10 % C10 AO in order to ensure the induction of local irritation. This treatment resulted in a moderate and confluent erythema in all test group animals 48 or 72 hours after start of exposure. Challenge with 2 mL of a 0.04 % solution of C10 AO revealed no skin irritation in any animal.

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the present test conditions N,N-dimethyldecylamine-N-oxide revealed no sensitising properties in guinea pigs in a test model according to MAGNUSSON and KLIGMAN.

Executive summary:

The skin sensitisation potential of C10 AO was assessed in a study performed according to OECD TG 406 (Magnusson and Kligman Maximisation Test) under GLP using male Dunkin-Hartley Guinea pigs.

A preliminary study was performed to determine the appropriate dose level of C10 AO to use in the main study following intracutaneous and topical administration. Six concentrations of C10 AO in aqueous solution were tested by intracutaneous injection using two animals. The first animal received doses (0.1 mL) of 0.4, 2 and 4 % C10 AO in aqueous solution. The second animal received doses (0.1 mL) of 0.004, 0.04 and 0.2 % C10 AO in aqueous solution. A concentration of 0.004% C10 AO revealed a discrete or patchy erythema, concentrations of 0.04% or 0.2% C10 AO revealed a moderate and confluent erythema 24 to 72 hours after administration, respectively. Concentrations of 0.4%, 2% or 4% C10 AO revealed an intense erythema and swelling with necrosis 24 to 72 hours after administration. A total of ten animals were used for topical administration. The test area of four animals was shaved, whilst for six animals the test area was shaved and then depilated. The animals were then treated occlusively with one or two concentrations of C10 AO solution (2 mL per application). The patches were removed after 24 hours (shaved and depilated) or 48 hours (shaved-only) and assessed immediately, 24 and 48 hours after removal (shaved and depilated) or immediately and 24 hours (shaved-only) after removal for erythema and oedema. The concentrations of C10 AO tested were 0.4, 2, 4, 10, 20, 30 and 40 % to the shaved-only and shaved and depilated skin and in addition 0.004, 0.02, 0.04 and 0.2% to the shaved and depilated skin. In the shaved-only animals concentrations of 0.4% to 10% revealed a moderate and confluent erythema 24 hours and a discrete or patchy erythema 48 hours after removal of the patches. The animals that received the 20% + 30% or 40% concentrations (equivalent to 3,300 and 2,640 mg/kg respectively) died prematurely. The cause of death could not be confirmed in this study. In the shaved and depilated animals no skin reactions were observed up to the concentration of 0.04%. The concentration of 0.2% revealed a discrete or patchy erythema, concentrations of 0.4% or 2% revealed a moderate and confluent erythema 24 to 72 hours after start of exposure, respectively. Concentrations of 4% or 10% revealed a moderate and confluent erythema with brownish-green spots on the skin 24 to 72 hours after start of exposure. The animals receiving the 20% + 30% or 40% concentration (equivalent to 3,300 and 2,640 mg/kg respectively) died prematurely. The cause of death could not be confirmed in this study. In a subsequent study performed according to OECD TG 402 (see section 7.2.3) the acute dermal toxicity to rats was determined to be >2000 mg AO/kg bw, with no deaths or signs of toxicity observed in the study. Based on the results of the preliminary study, it was decided to use a 0.2% concentration in tap water for the 1st (intracutaneous) induction stage, a 10% concentration in tap water for the 2nd (topical) induction stage and a 0.04% solution in tap water for the challenge in the main study. The main study was performed using 20 animals and a further 10 animals were used as vehicle (tap water) controls. For the intradermal induction the animals received three pairs of injections (0.1 mL each) into the shoulder region as follows: Freund’s complete adjuvant (diluted 1:1 with 0.9 % NaCl); 0.2 % C10 AO solution; 0.2 % C10 AO solution in a 1:1 mixture with FCA/0.9 % NaCl solution. On Day 6, prior to topical induction, the shoulder area was shaved again and the skin coated within sodium lauryl sulfate (0.5 mL, 10 % solution in vaseline) in order to ensure the induction of local irritation. Although this step is considered unnecessary based on the irritation seen in the preliminary study, it was standard practice in the laboratory that conducted the study. On Day 7 the animals were exposed topically under occlusive conditions to a 10 % C10 AO solution (2 mL) for 48 hours.The test animals and control animals were challenged with a 0.04% solution of C10 AO two weeks after the final induction. The test substance was applied to the shaved and depilated flank of the test animal and covered occlusively for 24 hours. Dermal reaction to the challenge was assessed 24 and 48 hours after the challenge exposure. No positive responses were noted at 24 or 48 hours.