Allyl heptanoate

Endpoint summary

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:

2010-01-14 to 2010-03-27

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP guideline study reliable with restrictions due to skin irritation effects caused by challenge concentration in all animals.

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

, adopted 1992-07-17

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

signed 2009-11-12

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

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

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland GmbH, Research Models and Services, Germany GmbH, Stolzenseeweg 32-36, 88353 Kißlegg, Germany
- Age at study initiation: 39 days
- Weight at study initiation: 328 - 372 g (excluding positive control group); Positive control group: 319 - 346 g
- Housing: The animals were kept in pairs in MAKROLON cages (MZK 80/25). Granulated textured wood (Granulat A2, J. BRANDENBURG, 49424 Goldenstedt, Germany) was used as bedding material in the cages.
- Diet (ad libitum): Commercial diet, ssniff® Ms-H V2233 (ssniff Spezialdiäten GmbH, 59494 Soest, Germany)
- Water (ad libitum): Tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 3 °C (maximum range)
- Relative humidity: 55% ± 15% (maximum range)
- Photoperiod (hrs dark / hrs light): 12/12
No further information on the test animals was stated.

Route:

intradermal and epicutaneous

Vehicle:

other: sesame oil

Concentration / amount:

Intracutaneous (induction): 5% solution of Allyl heptanoate (Sym09/611041) in sesame oil
Topical/shaved skin (induction): 50% solution of Allyl heptanoate (Sym09/611041) in sesame oil
Topical/shaved skin (challenge): 1% solution of Allyl heptanoate (Sym09/611041) in sesame oil

Route:

epicutaneous, occlusive

Vehicle:

other: sesame oil

Concentration / amount:

Intracutaneous (induction): 5% solution of Allyl heptanoate (Sym09/611041) in sesame oil
Topical/shaved skin (induction): 50% solution of Allyl heptanoate (Sym09/611041) in sesame oil
Topical/shaved skin (challenge): 1% solution of Allyl heptanoate (Sym09/611041) in sesame oil

No. of animals per dose:

Preliminary test: 20 animals
Main study: 20 animals (treatment group) plus 10 animals (vehicle control group)
The positive control group that was tested concurrently included 20 animals.

Details on study design:

RANGE FINDING TESTS:
The aim of the preliminary study was to determine the appropriate dose level of the test item following intracutaneous and topical administration. The concentration used in the main study following the induction procedure should produce mild to moderate skin (grade1 or grade 2) reactions following topical application and be adjusted to the highest level that can be well tolerated locally and generally following intracutaneous injection. For the challenge exposure a subirritating concentration was used which produced no evidence of skin irritation in non-sensitised animals.
The shoulder and the flank region of the animals were shaved or shaved and depilated (approx. 5 x 5 cm^2).
(a) intracutaneous: 0.1 mL of the prepared test item was administered intracutaneously (shoulder region).
Three concentrations of the test item were injected intradermally into two animals each, 3 further concentrations into two further animals.
(b) topical: The test area of 16 animals each was shaved or shaved and depilated. 2 mL of the test preparation was spread over a filter paper (2 x 4 cm) and applied to the test area and held in contact by an occlusive dressing .
Two concentrations each were applied to the shaved or shaved and depilated flanks of 16 animals each.
The occlusive dressing and the filter paper containing the test item were removed after 24 or 48 hours. No cleaning of the application site was necessary as the test item was completely absorbed. The application sites were assessed immediately, 24 and 48 (depilated) or immediately and 24 hours (non-depilated) after removal of the filter paper for erythema and oedema.
A 10 % (w/v) solution of Allyl heptanoate (Sym09/611041) in sesame oil was the maximum technically feasible concentration that could be administered by intradermal injection, since higher concentrations blocked that syringe and did not allow proper intracutaneous administration.

Results preliminary test:
Six concentrations of Allyl heptanoate (Sym09/611041) in sesame oil were tested by intracutaneous injection: 0.01, 0.1, 0.5, 1, 5 or 10% solutions in sesame oil: No skin reactions were observed up to the concentration of 0.5%. Concentrations of 1% to 5% revealed a discrete or patchy erythema, a concentration of 10% revealed a moderate and confluent erythemya, 24 to 72 hours after start of administration, respectively.
Several concentrations of Allyl heptanoate (Sym09/611041) in sesame oil were tested by topical application:
Undepilated skin:
Five concentrations of Allyl heptanoate (Sym09/611041) in sesame oil were tested: 1, 5, 10, 25 and 50% solutions in sesame oil and the undiluted test item.
No skin reactions were observed at the concentration of 1%. Concentrations of 5%, 25% and 50% revealed a discrete or patchy erythema in 2 of 2 animals, a 10% concentration in 1 of 2 animals, 48 and 72 hours after start of exposure, respectively. The undiluted test item revealed a moderate and confluent erythema in 2 of 2 animals 48 and 72 hours after start of exposure.
Depilated skin:
Nine concentrations of Allyl heptanoate (Sym09/611041) in sesame oil were tested: 0.01, 0.05, 0.1, 0.5, 1, 5, 10, 25 and 50% solutions in sesame oil and the undiluted test item.
Concentrations of up to 0.05% did not reveal any skin reaction. Concentrations of 0.1% to 25% revealed a discrete or patchy erythema in 2 of 2 animals 48 and 72 hours or 24 to 72 hours after start of exposure. The 2 animals treated with the 50% concentration and the undiluted test item died prematurely.
Due to the extreme test item skin reaction on depilated skin up to a concentration of 0.05% it was decided using a 1% concentration without depilation (only shaved) for the challenge.
The concentration used for the 1st (intracutaneous) induction stage was 5%, and 50% concentration for the 2nd (topical) induction stage.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 (Intradermal injection and topical administration)
- Test groups:
Day 0: 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) Freud's complete adjuvant (batch No. 049 K8700; SIGMA-ALDRICH Chemie GmbH, 82024 Taufkirchen, Germany)(diluted 1:1 with 0.9 % NaCl (batch No. 912816; Delta Select GmbH, 63303 Dreieich, Germany))
(2) the test item (5% in sesame oil)
(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 in injection 2.
Day 6:
As Allyl heptanoate (Sym09/611041) was only slightly irritating to the non-depilated skin of the tet animals in the preliminary experiment, the fur was shaved from the application area and the exposed skin was coated with 0.5 mL sodium laurylsulfate 10 % (batch no. 121K0370; SIGMA Chemie GmbH, 89420 Höchstädt, Germany) in vaseline in order to induce a local irritation.
Day 7:
7 days after the intracutaneous injection, the shoulder region of the same animals was shaved again but not depilated and treated topically using the patch-test technique (exposure period: 48 hours). No cleaning of the skin was necessary.
- Control group: The vehicle control animals were treated in the same was as the animals of the test group (2), but received sesame oil instead of the test item.
- Skin obervation and scoring: The skin reaction of the first induction exposure were evaluated at 24 and 48 hours, of the second induction at 48 and 72 hours after beginning of exposure.

B. CHALLENGE EXPOSURE
- No. of exposures: 1 (topical application)
- Test groups: Two weeks after the topical application (corrsponds to a monitoring period of 21 days) the flanks of the same animals were shaved but not depliated for a further topical application using the patch-test technique. The filter paper containing th test item was applied to the left flank, the filter with the vehicle to the right flank of the animal (exposure time: 24 hours). 21 hours after te filter paper had been removed, no cleaning of the treated skin was necessary.
- Control group: The left flank was treated with the test item, the right flank with the vehicle i.e. in the same way as the test group (2).
- Evaluation (hr after challenge):
Days 23 and 24: 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.The skin reactions were graded following the grading scale of MAGNUSSON/KLIGMAN. For evaluation, the percentage frequency and the degree of sensitisation only of those animals were used which exhibited irritation after the challenge (48-h value).

Other observations:
- Mortality: daily during the observation period
- Clinical signs: daily during the observation period
- Body weight: at start of study and at study termination
- Pathology: No necropsy was performed

Positive control group: Benzocaine solution (Benzocaine was dissolved in 40 % ethanolic 0.9% NaCl solution for phase 1, 2 and 3.): The animals of the positive control group (same origin (strain) as those used in the study) were treated with a 2% (w/v) benzocaine solution intracutaneously (2 X 0.1 mL/animal) in stage 1, a 5% (w/v) benzocaine solution topically (2 mL/animal) in stage 2 and stage 3. The aimals were shaved and not depliated in all stages.
No further information on the study design was stated.

Challenge controls:

Vehicle control group: sesame oil
10 males were used for the vehicle control group.

Positive control substance(s):

yes

Remarks:

benzocaine

Positive control results:

Animals of the same strain treated with benzocaine in 40% ethanolic 0.9% NaCl solution exhibited a sensitising reaction in all animals in form of a moderate and confluent erythema or an intense erythema and swelling.

Reading:

1st reading

Hours after challenge:

48

Group:

positive control

Dose level:

5% (w/v) benzocaine solution topically

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

all animals exhibited a sensitising reaction in form of a moderate and
confluent erythema (grade 2) or an intense erythema and swelling (grade 3)

Remarks on result:

other: 48 hours after starting of the challenge

Reading:

2nd reading

Hours after challenge:

72

Group:

positive control

Dose level:

5% (w/v) benzocaine solution topically

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

all animals exhibited a sensitising reaction in form of a moderate and
confluent erythema (grade 2) or an intense erythema and swelling (grade 3)

Remarks on result:

other: 72 hours after starting of the challenge

Reading:

1st reading

Hours after challenge:

48

Group:

test chemical

Dose level:

1% solution of Allyl heptanoate (Sym09/611041) in sesame oil

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

In all animals discrete or patchy erythema was observed.

Remarks on result:

other: 48 hours after starting of the challenge. Same effects were observed both in control and test item animals. Hence, the observed effects are not considered as sensitising effects.

Reading:

2nd reading

Hours after challenge:

72

Group:

test chemical

Dose level:

1% solution of Allyl heptanoate (Sym09/611041) in sesame oil

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

In all animals discrete or patchy erythema was observed.

Remarks on result:

other: 72 hours after starting of the challenge. Same effects were observed both in control and test item animals. Hence, the observed effects are not considered as sensitising effects.

Reading:

1st reading

Hours after challenge:

48

Group:

negative control

Dose level:

1% solution of Allyl heptanoate (Sym09/611041) in sesame oil; no test item during the induction stages

No. with + reactions:

10

Total no. in group:

10

Clinical observations:

In all animals discrete or patchy erythema was observed.

Remarks on result:

other: 48 hours after starting of the challenge. Same effects were observed both in control and test item animals. Hence, the observed effects are not considered as sensitising effects.

Reading:

1st reading

Hours after challenge:

72

Group:

negative control

Dose level:

1% solution of Allyl heptanoate (Sym09/611041) in sesame oil; no test item during the induction stages

No. with + reactions:

10

Total no. in group:

10

Clinical observations:

In all animals discrete or patchy erythema was observed.

Remarks on result:

other: 72 hours after starting of the challenge. Same effects were observed both in control and test item animals. Hence, the observed effects are not considered as sensitising effects.

Discrete or patchy erythema did not reveal only in those animals which were treated with test item during the previous stages but also in those of the vehicle control. In the challenge, the left flank of the control animals was also treated with the test item (the right flank with the vehicle) to exclude sensitising properties of the test item due to irritating effects.

Hence, the effects observed in the animals of the control and treated animals are considered to be due to a marginal irritating effect of the test item, but not a sensitising effect.

The body weight gain of the animals treated with Allyl heptanoate (Sym09/611041) was within the range of the vehicle control during the experiment (48.6% versus 49.2% (mean values).

Behaviour of the animals remained unchanged.

Interpretation of results:

not sensitising

Remarks:

Migrated information

Conclusions:

Under the present test conditions Allyl heptanoate revealed no sensitising properties in guinea pigs in a test model according to MAGNUSSON and KLIGMAN. Therefore, the test item must not be classified and labelled according to regulation (EC) No. 1272/2008.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Under the present test conditions Allyl heptanoate revealed no sensitising properties in guinea pigs in a test model according to Magnusson and Kligman.

Migrated from Short description of key information:
No sensitisation effects were noted.

Justification for selection of skin sensitisation endpoint:
Reliable in vivo study using maximisation test

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Skin Sensitisation :

Under the present test conditions Allyl Heptanoate revealed no sensitising properties in guinea pigs in a test model according to Magnusson and Kligman, OECD 406. Hence, classification according to Regulation (EC) No 1272/2008 or Directive 67/548/EEC is not required.