Undec-10-enoic acid

Skin sensitisation

Administrative data

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:

guideline study

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.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

July 1992

Deviations:

no

Test guideline 2

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

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.

Test material

Test material information

Constituent 1

Reference substance name:

Undec-10-enoic acid

EC Number:

203-965-8

EC Name:

Undec-10-enoic acid

Cas Number:

112-38-9

Molecular formula:

C11H20O2

IUPAC Name:

undec-10-enoic acid

Constituent 2

Reference substance name:

Undecylenic acid

IUPAC Name:

Undecylenic acid

Test material form:

other:

Details on test material:

- Name of test material: Undecylenic acid
- Description at liquefactiona: translucent and homogenous liquid
- CAS number : 112-38-9
- Lot/batch No.: 1307020
- Analytical purity: 98.56%
- Expiry date: 24 March 2015
- Storage condition: at room temperature

In vivo test system

Test animals

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

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

corn oil

Induction 2

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

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

corn oil

Challenge 2

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

Results and discussion

Positive control results:

no positive control

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

other: 3 reading

Hours after challenge:

72

Group:

test chemical

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

Results 2

Key result

Reading:

other:

Hours after challenge:

72

Group:

negative control

Dose level:

0

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

very slight erythema

Remarks on result:

other: negative

Any other information on results incl. tables

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.

 

 

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

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.