5-methyl-5-phenylhexan-3-one

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The study was conducted between 13 September 2004 and 29 November 2004.

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

In accordance with GLP conditions, but tested only up to 30%

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2004

Report date:

2004

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

yes

Remarks:

maximum concentration tested was 30%

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

5-methyl-5-phenylhexan-3-one

EC Number:

225-561-0

EC Name:

5-methyl-5-phenylhexan-3-one

Cas Number:

4927-36-0

Molecular formula:

C13H18O

IUPAC Name:

5-methyl-5-phenylhexan-3-one

Test material form:

liquid

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/J

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Jackson Laboratories, Bar Harbor, ME 04609, US
- Age at study initiation: 11 weeks at start of dosing
- Weight at study initiation: 18 to 26 g at the outset of the study (Day 1)
- Housing: 5 animals/cage (randomly allocated)
- Diet (e.g. ad libitum): ad libitum (Certified Rodent Chow 7012C)
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.1-26.7
- Humidity (%): 30-53
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

other: Diethyl phthalate/ Ethanol 3:1

Concentration:

7.5%, 15% and 30% (w/v)

No. of animals per dose:

5

Details on study design:

MAIN STUDY

EXPERIMENTAL PHASE
Groups of five mice were treated with the test item at concentrations of 7.5%, 15% and 30% w/v in DEP/EtOH. No justification on the choice of vehicle is provided. The doses were chosen on the basis that these are typical concentrations used for fragrances. The mice were treated by daily application of 25 µl of the appropriate concentration of the test item to the dorsal surface of both ears for three consecutive days (Days 1, 2, 3). The test item formulation was administered using an automatic micropipette and spread over the dorsal surface of the ear using the tip of the pipette.
A further group of five mice received the vehicle alone in the same manner. The positive control animals were similarly treated to the test animals except that 25 µl of the positive control item, Hexylcinnamaldehyde [HCA], at a concentration of 35% v/v. On day 6, all mice were injected i.v. with 250 µl of sterile saline containing 20µCi of 3H-thymidine.

OBSERVATIONS
- Clinical Observations: All animals were observed before dosing and once post-dose on Day 1 to 3. After this observations continued on a daily basis on Days 4, 5 and 6. Any signs of toxicity or signs of ill health during the test were recorded. The animals were also examined daily for erythema and edema on the site of application.
- Bodyweights: The bodyweight of each mouse was recorded on Day 1 (prior to dosing) and Day 6 (prior to termination).

TERMINATIOIN
Five hours following the administration of 3HTdR all mice were killed by carbon dioxide asphyxiation. For each individual animal of each group the draining auricular lymph nodes were excised and processed. A single cell suspension of the lymph node cells for each individual animal was prepared. The lymph node cells were rinsed with PBS and precipitated with 5% trichloroacetic acid during night. The lymph node cells suspension was transferred to a centrifuge tube and thereafter, the pellets were resuspended in 1 ml TCA and transferred to scintillation fluid vials for the measurement of the radioactive material.

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Statistics:

Data was processed to give group mean values for disintegrations per minute and standard deviations where appropriate. Increases in the radioactivity compared to the vehicle control were recorded as stimulation indices (SI). Individual DPM values were analyzed with log values. Dunett's test was used to determine significance when required.

Results and discussion

Positive control results:

The positive control item, Hexylcinnamaldehyde, gave a Stimulation Index of 7.11 which was statistically significant when compared to the vehicle control group.

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

EC3

Test group / Remarks:

All test groups

Remarks on result:

other: up to 30% alll SI results are < 3

Results 2

Parameter:

SI

Value:

0.89

Test group / Remarks:

7.5% (w/v)

Results 3

Parameter:

SI

Value:

0.8

Test group / Remarks:

15% (w/v)

Results 4

Parameter:

SI

Value:

1.12

Test group / Remarks:

30% (w/v)

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
Mean DPM (+/- SEM) were:
- Vehicle control: 233 +/- 44
- 7.5%: 207 +/- 39
- 15%: 187 +/- 18
- 30%: 260 +/- 7

DETAILS ON STIMULATION INDEX CALCULATION
Stimulation Index = dpm/lymph node test item treated group / dpm/lymph node vehicle control.
- 7.5%: SI = 0.89
- 15%: SI = 0.8
- 30%: SI = 1.12

EC3 CALCULATION
A dose-response relationship was not observed. A calculation of the EC3 value was not performed because no test concentrations produced a S.I. of 3 or higher.

CLINICAL OBSERVATIONS:
One mouse in the group treated with the test article at 15% was found dead on day 5. A necropsy revealed no gross lesions. All other mice survived and did not show signs of toxicity. The lymph nodes of all animals which were treated with the test item were normal in size and appearance. At termination both ears were measured. There was one statistically significant different in the ear measurement in the group treated with the test article at a dose of 7.5%. This slight decrease was not considered to be biologically significant.

BODY WEIGHTS
Body weight changes of the test animals between Day 1 and Day 6 were comparable to those observed in the corresponding control group animals over the same period.

Applicant's summary and conclusion

Interpretation of results:

other: not sensitising

Remarks:

based on CLP criteria (EC 1272/2008 and its updates)

Conclusions:

Under the conditions of this test, Damascol did not elicit a Stimulation Index of greater than 3 and was therefore not considered to be a sensitiser up to 30%. Therefore, the substance does not need to be classified as skin sensitiser in accordance with the criteria outlined in Annex I of the CLP Regulation (1272/2008/EC and its updates).

Executive summary:

The skin sensitisation potential of the test substance has been tested according to OECD TG 429 (Local Lymph Node Assay) and under GLP conditions. The positive control showed a statistically significant induction of the stimulation index as compared to the vehicle control. At doses of 7.5, 15 and 30% (w/v) Damascol, the following Stimulation Index (SI) values were found, respectively: 0.89, 0.80 and 1.12. As these SI values are all below 3 and no EC3 value could be calculated, the substance was not considered to be a skin sensitiser under the conditions of this test. Therefore, the substance does not need to be classified as skin sensitiser up to 30%.