Registration Dossier

Diss Factsheets

Toxicological information

Skin sensitisation

Currently viewing:

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2012-2013
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: The study was conducted according to GLP and valid testing guidelines, therefore it is considered relevant, adequate and reliable for classification.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of study:
guinea pig maximisation test

Test material

Constituent 1
Reference substance name:
Butanoic acid, 4-amino-4-oxo-2(or 3)-sulfo-, N-(C16-C18 (even numbered), C18 unsaturated alkyl), disodium salts
IUPAC Name:
Butanoic acid, 4-amino-4-oxo-2(or 3)-sulfo-, N-(C16-C18 (even numbered), C18 unsaturated alkyl), disodium salts
Test material form:
other: Liquid to pasty
Details on test material:
- Name of test material (as cited in study report): Butanoic acid, 4-amino-4-oxo-2(or3)-sulfo-, N-(C16-C18 (even numbered), C18unsaturated alkyl)), disodium salts; Sodium tallow sulphosuccinamates
- Physical state: Liquid to pasty; opaque
- Analytical purity: 25.5% active matter (correction factor: Not applicable)
- Impurities (identity and concentrations): See confidential details
- Composition of test material, percentage of components: See confidential details
- Purity test date: February 14, 2012
- Lot/batch No.: MIBX042332
- Expiration date of the lot/batch: February 14, 2013
- Stability under test conditions: stable
- Storage condition of test material: At +10° to +25 °C
- Other: Manufacturer/supplier: Huntsman Holland BV, Merseyweg 10, 3197 KG Botlek-Rotterdam, The Netherlands

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 Deutschland GmbH, Stolzenseeweg 32 – 36, 88353 Kisslegg, Germany
- Age at start of administration: Approx. 5 weeks
- Weight at start of administration: 322 – 372 g (excluding positive control group); 316 - 370 g (positive control group)
- Housing: In pairs in MAKROLON cages (MZK 80/25). Granulated textured wood (Granulat A2, J. Brandenburg, 49424 Goldenstedt, Germany) was used as bedding material for the cages. The cages were changed and cleaned twice a week.
- Diet (e.g. ad libitum): Commercial diet, ssniff® Ms-H (ssniff Spezialdiäten GmbH, 59494 Soest, Germany) ad libitum.
- Water (e.g. ad libitum): Drinking water was offered ad libitum and examined according to the "Deutsche Trinkwasserverordnung 2001 " by the Hamburger Wasserwerke, 20539 Hamburg, Germany, at least four times a year. In addition, drinking water samples taken at LPT are analysed by LUFA-ITL once a year for means of bacteriological investigations according to the "Deutsche Trinkwasserverordnung 2001, Anlage 1 ".
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C ± 3°C (maximum range)
- Humidity (%): 55% ± 15% (maximum range).
- Air changes (per hr): Not provided
- Photoperiod (hrs dark / hrs light): 12/12

IN LIFE DATES: From: October 29, 2012 To: February 9, 2013

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
other: lntracutaneous(stage 1) and Topical (stage 2)
Vehicle:
other: Tap water
Concentration / amount:
Preliminary (Range Finding) test:
Stage1: intracutaneous: 10, 5, 1, 0.5, 0.1 and 0.01% concentration of the test item in tap water
Stage 2: topical: 100, 75, 50, 25, 10, 5 and 1% concentration of the test item in tap water
Stage 3: topical: 100, 75, 50, 25, 10, 5 and 1% concentration of the test item in tap water


Main test:
Stage 1: intracutaneous: 0.1% suspension of the test item in tap water
Stage 2: topical: undiluted test item
Stage 3: topical: 10% suspension of the test item in tap water
Challengeopen allclose all
Route:
other: Topical (stage 3)
Vehicle:
other: Tap water
Concentration / amount:
Preliminary (Range Finding) test:
Stage1: intracutaneous: 10, 5, 1, 0.5, 0.1 and 0.01% concentration of the test item in tap water
Stage 2: topical: 100, 75, 50, 25, 10, 5 and 1% concentration of the test item in tap water
Stage 3: topical: 100, 75, 50, 25, 10, 5 and 1% concentration of the test item in tap water


Main test:
Stage 1: intracutaneous: 0.1% suspension of the test item in tap water
Stage 2: topical: undiluted test item
Stage 3: topical: 10% suspension of the test item in tap water
No. of animals per dose:
Preliminary test: 10 animals in total ( 8 topical, 2 intracutaneous administration)
Main test: 5 animals (vehicle control), 10 animals (treatment), 20 animals (positive control: α-hexyl cinnamic aldehyde: historical data, not tested concurrently )
Details on study design:
RANGE FINDING TESTS:
Six concentrations of Butanoic acid, 4-amino-4-oxo-2(or 3)-sulfo-, N-(C16-C18 (even numbered), C18 unsaturated alkyl), disodium salts were tested by intracutaneous injection: 0.01, 0.1, 0.5, 1, 5 or 10% suspensions in tap water: A concentration of 0.01% revealed a discrete or patchy erythema 24 and 48 hours after administration. Concentrations of 0.1 or 0.5% revealed a moderate and confluent erythema 24 and 48 hours and a discrete or patchy erythema 72 hours after administration. Concentrations of 1%, 5% or 10% revealed an intense erythema and swelling 24, 48 and 72 hours after administration.
Seven concentrations of the test item were tested by topical application: 1, 5, 10, 25, 50 and 75% suspensions in tap water and the undiluted test item.
Non-depilated skin
Concentrations of 1%, 5% or 10% did not reveal any skin reaction. A concentration of 25% revealed a discrete or patchy erythema, a concentration of 50% revealed a moderate and confluent erythema 48 hours after start of exposure, respectively. A concentration of 75% or the undiluted test item revealed a moderate and confluent erythema 48 hours and a discrete or patchy erythema 72 hours after start of exposure.
Depilated skin
Concentration of 1%, 5% or 10% did not reveal any skin reaction. Concentrations of 25% or 50% revealed a discrete or patchy erythema 24 and 48 hours, the concentration of 75% 24, 48 and 72 hours after start of exposure. The undiluted test item revealed a moderate and confluent erythema 24 and 48 hours and a discrete or patchy erythema 72 hours after start of exposure.
Hence, it was decided, after consultation with the Sponsor, to use a 0.1% concentration in tap water for the 1st (intracutaneous) stage, the undiluted test item for the 2nd (topical) induction stage and a 10% suspension in tap water for the challenge.

MAIN STUDY
A. INDUCTION EXPOSURE

stage 1: Intradermal injections Day 0
- No. of exposures: 3 (pairs of injections)
- Exposure period: Single injections
- Test groups: 10 animals, 0.1% concentration test item in tap water
Three pairs of intradermal injections of 0.1 mL were given in the shoulder region which was cleared of hair so that one of each pair lay on each side of midline.
(1) Freund's complete adjuvant
(diluted 1 : 1 with 0.9% NaCl )
(2) the test item (concentration see above)
(3) the test item in a 1:1 mixture (v/v) FCA/physiological saline
In injection 3, the final concentration of the test item was equal to that used in injection 2.
Injections (1) and (2) were given close to each other and nearest the head, while (3) was given towards the caudal part of the test area.
- Control group:
Vehicle control: 5 animals:
(1)Freund's complete adjuvant (diluted 1 : 1 with 0.9% NaCl )
(2) tap water
(3) tap water in a 1:1 mixture (v/v) FCA/physiological saline
Positive control: 20 animals (historical data): 10% (v/v) α-hexyl cinnamic aldehyde solution
- Site: Shoulder region
- Frequency of applications: Single injections
- Duration: Single injections (Day 0) and six days interval before stage 2 (Day 7)
- Concentrations: 0.1% concentration test item in tap water

Stage 2: Topical Day 7
- No. of exposures: 1 (patch)
- Exposure period: 48 hours
- Test groups: undiluted test item
- Control group:
Vehicle control: 5 animals: tap water
Positive control: 20 animals (historical data): undiluted α-hexyl cinnamic aldehyde
- Site: Shoulder region
- Frequency of applications: 1
- Duration: 48 hours
- Concentrations: undiluted test item


B. CHALLENGE EXPOSURE
Stage 3: Topical Day 21
- No. of exposures: 1 (patch)
- Day(s) of challenge: Day 21
- Exposure period: 24 hours
- Test groups: The filter paper containing the test item (10% suspension of the test item in tap water) was applied to the left flank, the filter paper with the vehicle to the right flank of the animal.
- Control group:
Vehicle control: The left flank was treated with the test item (10% concentration in tap water), the right flank with the vehicle i.e. in the same way as the test group.
Positive control: 0.01% α-hexyl cinnamic aldehyde solution.
- Site: Flanks
- Concentrations: 10% suspension of the test item in tap water
- Evaluation (hr after challenge):
21 hours after removing the filter paper the challenge area was cleaned and cleared of hair if necessary; three hours later (at 48 hours from the start of challenge application) the skin reaction was observed and recorded; 24 hours after this observation a second observation (72 hours) was made and recorded.

OTHER:
MAGNUSSON/KLIGMAN grading scale tor the evaluation of challenge patch test reactions
0 = no visible change
1 = discrete or patchy erythema
2 = moderate and confluent erythema
3 = intense erythema and swelling

Challenge controls:
- Vehicle control: The left flank was treated with the test item (10% concentration in tap water), the right flank with the vehicle i.e. in the same way as the test group.
- Positive control: The filter paper containing 0.01% α-hexyl cinnamic aldehyde solution was applied to the left flank, the filter paper with the vehicle to the right flank of the animal.
Positive control substance(s):
yes
Remarks:
α-hexyl cinnamic aldehyde

Results and discussion

Positive control results:
Animals of the same strain treated with α-hexyl cinnamic aldehyde in sesame oil exhibited a sensitising reaction in all animals in form of a moderate and confluent erythema (grade 2).

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
48
Group:
test chemical
Dose level:
10% in tap water (2mL)
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 48.0. Group: test group. Dose level: 10% in tap water (2mL). No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
72
Group:
test chemical
Dose level:
10% in tap water (2mL)
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 72.0. Group: test group. Dose level: 10% in tap water (2mL). No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
1st reading
Hours after challenge:
48
Group:
negative control
Dose level:
tap water (2mL)
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 48.0. Group: negative control. Dose level: tap water (2mL). No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
2nd reading
Hours after challenge:
72
Group:
negative control
Dose level:
tap water (2mL)
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 72.0. Group: negative control. Dose level: tap water (2mL). No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
1st reading
Hours after challenge:
48
Group:
positive control
Dose level:
0.01% α-hexyl cinnamic aldehyde solution
No. with + reactions:
20
Total no. in group:
20
Clinical observations:
moderate and confluent erythema left flank
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 48.0. Group: positive control. Dose level: 0.01% α-hexyl cinnamic aldehyde solution. No with. + reactions: 20.0. Total no. in groups: 20.0. Clinical observations: moderate and confluent erythema left flank.
Reading:
2nd reading
Hours after challenge:
72
Group:
positive control
Dose level:
0.01% α-hexyl cinnamic aldehyde solution.
No. with + reactions:
20
Total no. in group:
20
Clinical observations:
moderate and confluent erythema left flank
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 72.0. Group: positive control. Dose level: 0.01% α-hexyl cinnamic aldehyde solution.. No with. + reactions: 20.0. Total no. in groups: 20.0. Clinical observations: moderate and confluent erythema left flank.

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, test item containing 25.5% was found to be not sensitising to guinea pigs in a test model according to MAGNUSSON and KLIGMAN.
Executive summary:

The purpose of this study was to determine the potential of test item, containing 25.5% active ingredient, to produce skin sensitisation reactions in guinea pigs in a test model according to MAGNUSSON and KLIGMAN.

Based on the preliminary study, it was decided, to use a 0.1% concentration in tap water for the 1st (intracutaneous) stage, the undiluted test item for the 2nd (topical) induction stage and a 10% suspension in tap water for the challenge.

A 0.1% suspension in tap water chosen for the 1st (intracutaneous) induction stage revealed a discrete or patchy erythema in all 10 animals 24 and 48 hours after administration. 2 mL of the undiluted test item/animal chosen for the 2nd (topical) induction stage revealed a moderate and confluent erythema in all 10 animals 48 and 72 hours after administration. The challenge with 2 mL of a 10% suspension of the test item in tap water/animal - the maximum non-irritating concentration - revealed no skin irritation in any animal and, thus, the test item had no sensitising properties.

The vehicle control revealed no skin reactions. Animals of the same strain treated with α-hexyl cinnamic aldehyde in sesame oil exhibited a sensitising reaction in all animals in form of a moderate and confluent erythema (grade 2).

Behaviour of the animals remained unchanged. Body weights were not influcenced.

Under the present test conditions, Butanoic acid, 4-amino-4-oxo-2(or 3)-sulfo-, N-(C16-C18 (even numbered), C18 unsaturated alkyl), disodium salts was found to be not sensitising to guinea pigs in a test model according to MAGNUSSON and KLIGMAN.