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Description of key information

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
05 March 2014 -- 01 July 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Compliant to GLP and testing guidelines; adequate consistence between data, comments and conclusions.
Justification for type of information:
A Magnusson and Kligman was performed for this undecylenic acid as it is known that this substance may induce false positive in the LLNA test.
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
A Magnusson and Kligman was performed for this undecylenic acid as it is known that this substance may induce false positive in the LLNA test.
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: breeder: Charles River Laboratories France, L’Arbresle, France.
- Age at study initiation: approximately 1 to 2 months old on the day of treatment
- Mean body weight at study initiation: the males had a mean body weight of 332 g (range: 300 g to 371 g) and the females had a mean body weight of 318 g (range: 291 g to 360 g).
- Fasting period before study: no
- Housing: polycarbonate cages with stainless steel lids
- Diet: R14C pelleted diet (free access)
- Water: tap water filtered with a 0.22 µm filter (free access)
- Acclimation period: at least 5 days before the beginning of the study

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): approximately 12 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12 h/12 h

IN-LIFE DATES: 25 March 2014 to 01 July 2014
Route:
intradermal and epicutaneous
Vehicle:
corn oil
Concentration / amount:
Induction phase:
Concentration for intradermal injection: 1% *
Concentration for topical application: 100% *
*: highest to cause mild-to-moderate skin irritation based on preliminary assays

Challenge phase:
Concentration for topical application: 2.5%**
** highest non-irritant concentration based on preliminary assays
Route:
epicutaneous, occlusive
Vehicle:
corn oil
Concentration / amount:
Induction phase:
Concentration for intradermal injection: 1% *
Concentration for topical application: 100% *
*: highest to cause mild-to-moderate skin irritation based on preliminary assays

Challenge phase:
Concentration for topical application: 2.5%**
** highest non-irritant concentration based on preliminary assays
No. of animals per dose:
- preliminary test: 20 treated animals (10 males and 10 females)
- main test: 30 animals (15 males and 15 females)
Details on study design:
MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2
- Exposure period: topical induction: 48h
- Site: interscapular region
- Frequency of applications: once intradermal, once topical

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 3 weeks between injection (induction) and challenge
- Exposure period: 24h
- Site: right flank (test item) and left flank (vehicle)
- Evaluation (hr after challenge): 24, 48 and 72 h after removal of dressing
Challenge controls:
propylene glycol
Positive control substance(s):
not required
Remarks:
mercaptobenzothiazole tested in another study
Key result
Reading:
other: 3 reading
Hours after challenge:
72
Group:
test group
Dose level:
2.5%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Erythema were observed but only suggesting irritant effect
Remarks on result:
no indication of skin sensitisation

Skin reactions (incidence and severity of erythema and presence of edema) during the challenge application

 

Left flank exposed to

vehicle

(propylene glycol)

Right flank exposed to

test item

at 2.5%

24h

48h

72h

 

24h

48h

72h

Group 6 (control)

No erythema (grade 0)

0%

0%

0%

30%

10%

40%

Discrete erythema (grade 1)

0%

0%

0%

50%

50%

60%

Moderate erythema (grade 2)

0%

0%

0%

20%

40%

0%

Intense erythema (grade 3)

0%

0%

0%

0%

0%

0%

Group 7 (treated)

No erythema (grade 0)

0%

0%

0%

10%

25%

70%

Discrete erythema (grade 1)

0%

0%

0%

70% (+20)

50% (0)

30% (-30)

Moderate erythema (grade 2)

0%

0%

0%

10% (-10)

25% (-15)

0% (0)

Intense erythema (grade 3)

0%

0%

0%

10% (+10)

0% (0)

0% (0)

( ): difference from controls, % to take into account to classify the test item according to the CLP Regulation.

 

 

Interpretation of results:
other: not be classified as a skin sensitizer
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
Under the experimental conditions of this study, when compared to controls, the test item did not induce delayed contact hypersensitivity in guinea pigs.

Therefore, the test item should not be classified as a skin sensitizer according to the criteria of CLP Regulation.
Executive summary:

The objective of this study was to evaluate the potential of the test item to induce delayed contact hypersensitivity in guinea pigs.

This study was conducted in compliance with OECD Guideline No. 406 and the principles of Good Laboratory Practices.

 

Methods

A preliminary test of five groups using 2 animals/sex/group was first performed in order to determine the vehicle and the test item concentrations to be used in the main test.

In the main test, 30 guinea pigs (15 males and 15 females) were allocated to two groups: one control group of 5 males and 5 females (group 6) and one treated group of 10 males and 10 females (group 7).

 

On Day 1, three pairs of intradermal injections (0.1 mL/injection) were performed in the interscapular region of animals:

- Freund's Complete Adjuvant (FCA) diluted to 50% (v/v) with 0.9% NaCl,

- test item at the concentration of 1% in corn oil (treated group 7) or vehicle alone (control group 6),

- test item at the concentration of 1% in a mixture FCA/0.9% NaCl (50/50, w/w) (treated group 7) or vehicle at the concentration of 50% (w/v) in FCA/0.9% NaCl (50/50, v/v) (control group 6).

 

As in the preliminary test, the highest well-tolerated concentration of 100% was shown to be non-irritant after topical application, 0.5 mL of sodium lauryl sulfate at 10% (w/w) in vaseline was applied to the induction site on Day 7 in order to induce a local irritation.

On Day 8, a filter paper (approximately 8 cm2) was fully-loaded with the test item at the concentration of 100%, and then applied to the clipped interscapular region, over the intradermal injection sites of treated group 7 animals. The filter paper was held in place by an occlusive dressing for 48 hours. The control group 6 animals received an application of water for injection under the same experimental conditions. The presence of local irritation was checked (but not scored).

The induction phase was followed by a 14-day rest period.

 

On Day 22, all groups 6 and 7 animals were challenged by a cutaneous application of the test item at the concentration of 2.5% (w/w) in propylene glycol to the right flank. The Finn Chamber r filter paper was held in contact with the skin by an occlusive dressing for 24 hours. The vehicle was applied to the left flank under the same experimental conditions. Cutaneous reactions were evaluated before treatment, 24, 48 and 72 hours after removal of the dressing.

 

Each animal was observed at least once a day for mortality and clinical signs during the treatment and observation periods. Body weight was recorded on Day 1 and at the end of each observation period.

 

On completion of the observation period, the animals were sacrificed then discarded without macroscopic post-mortem examination. For all animals, skin samples of the challenged application sites were preserved. A microscopic examination was performed.

Results

According to the criteria for the selection of concentrations described in the OECD 406 test guideline and according to the results of solubility assays, the concentration selected for intradermal injections in the main test was 1% incorn oil,the concentration selected for the topical application of the induction phase (Day 8) was 100% and the concentration chosen for the challenge application (Day 22) was 2.5%. Propylene glycol was the vehicle retained for the challenge topical application.

Local reactions after challenge application:

Discrete erythema recorded on the right flank (exposed to test item) in treated group 7 was of lower or same incidence than that observed in control group 6 at 48- and 72-hour readings but was of higher incidence at 24-hour reading (70% vs. 50% erythema incidence in control group 6).

 

According to the criteria of CLP Regulation, the difference of 20% noted for discrete erythema was below the criteria of 30% to class the test item as a skin sensitizer.

Moderate erythema observed on the right flank (exposed to test item) in treated group 7 was of lower incidence than that noted in control group 6 at all reading times.

In addition, intense erythema observed on the right flank (exposed to the test item) of treated group 7 was only observed in a very few animals at 24-hour reading suggestingirritant effects of the test item.

The above described cutaneous reactions never exceeded the 30% classification threshold of the test item as skin sensitizer and were not considered to be related to contact delayed hypersensitivity. They rather suggested irritant effects of the test item despite the choice of a non-irritant concentration according to the results of the challenge phase in the preliminary test.

 

Therefore, the cutaneous reactions were attributed to the irritant properties of the test item.

 

Microscopic examination:

No signs of delayed hypersensitivity were observed with the test item or the vehicle.

Signs of local irritation were observed at both sites of application (vehicle and test item) in the control and treated groups. These changes were similar in nature and severity in both groups and consisted of epidermal changes (serocellular crusts, hyperkeratosis, acanthosis) and dermal changes (infiltrate of mixed inflammatory cells) which were seen at a higher incidence and/or severity at the sites treated with the test item than at this treated with the vehicle.

In addition, minimal or slight hemorrhage and edema were seen in the dermis of occasional test item-treated sites only.

The minimal isolated foci of spongiosis with exocytosis observed at a few sites including the test item-treated sites were not considered to represent signs of delayed hypersensitivity but were considered as part of the inflammation.

 

Discussion

As the cutaneous reactions observed in the animals of the treated group were of similar or lower incidence and severity when compared to those recorded in the animals of the control group, they were considered to be attributed to the irritant properties of the test item but not to delayed contact hypersensitivity.

 

Conclusion

Under the experimental conditions of this study, the test item did not induce delayed contact hypersensitivity in guinea pigs.

Therefore, the test item should not be classified as a skin sensitizer according to the criteria of CLP Regulation.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Two studeis are available for this endpoint. One OECD 406 performed with an analog susbtance, sodium undecylenate and a second GLP OECD 406 perfomed with the test item undecylenic acid.

A GLP study performed according to OECD 406 using undecylenic acid was perfomeed (Papineau, 2014). According to the criteria for the selection of concentrations described in the OECD 406 test guideline and according to the results of solubility assays, the concentration selected for intradermal injections in the main test was 1% incorn oil,the concentration selected for the topical application of the induction phase (Day 8) was 100% and the concentration chosen for the challenge application (Day 22) was 2.5%. Propylene glycol was the vehicle retained for the challenge topical application.

As the cutaneous reactions observed in the animals of the treated group were of similar or lower incidence and severity when compared to those recorded in the animals of the control group, they were considered to be attributed to the irritant properties of the test item but not to delayed contact hypersensitivity. It was therefore concluded that the test item did not induce delayed contact hypersensitivity in guinea pigs.

A second supportive study was performed with the analog, SODIUM UNDECYLATE (in a 33.2% aqueous solution), according to the maximization procedure by Magnusson and Kligman (Clauzeau, 1989). The test substance was applied by intradermal injection at a concentration of 0.1% in physiological saline, after 9 days (and irritation by sodium lauryl sulfate in vaseline) epicutanously at a concentration of 0,05%, and again for challenge at the latter concentration and route on a different section of the skin. No symptoms, mortality or cutaneaous reactions due to any sensitization process by treatment with the test substance were observed (0%).

 

Therefore, the test item should not be classified as a skin sensitizer according to the criteria of CLP Regulation.


Migrated from Short description of key information:
According to the OECD 406 guideline, a negative guinea pig maximization test was conducted in 1989 using sodium undecylenate. A second GLP
study performed on undecylenic acid according to OECD 406 confirmed that undecylenic acid is not a kin sensitizer (Papineau 2014).

Respiratory sensitisation

Endpoint conclusion
Additional information:
Migrated from Short description of key information:
no test requirement

Justification for classification or non-classification

According to EU regulation (EC) No 1272/2008 (CLP) and to EU Directive 67/584/EEC, undecylenic acid is not classified as a skin sensitizer.