4-(1-methoxy-1-methylethyl)-1-methylcyclohexene

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 June 2005 - 13 July 2005

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Justification for type of information:

Information from a reliable, Buehler test is available being performed before the change in the Reach regulation, preferring the in vitro tests.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

1992

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Version / remarks:

2003

GLP compliance:

yes

Type of study:

Buehler test

Justification for non-LLNA method:

This study is available and is sufficient to determine the skin sensitizing potential of the substance.

Test material

Test material information

1

Reference substance name:

4-(1-methoxy-1-methylethyl)-1-methylcyclohexene

EC Number:

238-620-0

EC Name:

4-(1-methoxy-1-methylethyl)-1-methylcyclohexene

Cas Number:

14576-08-0

Molecular formula:

C11H20O

IUPAC Name:

4-(2-methoxypropan-2-yl)-1-methylcyclohex-1-ene

Test material form:

liquid

In vivo test system

Test animals

Species:

guinea pig

Strain:

Hartley

Remarks:

Albino

Sex:

male/female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Elm Hill Breeding Laboratories, Chelmsford, MA
- Females nulliparous and non-pregnant: yes
- Age at study initiation: young adults
- Weight at study initiation:357 – 455 g
- Housing: in suspended stainless steel caging with mesh floors which confirm to the size recommendations of the Guide for the Care and Use of Laboratory Animals DHEW. Litter paper was placed beneath the cage and was changed at least three times per week.
- Diet: Pelleted Purina Guinea Pig Chow #5025
- Water: filtered tap water, ad libitum
- Acclimation period: 12 to 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17 - 23
- Humidity (%): 40 - 66
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Induction

Route:

epicutaneous, occlusive

Vehicle:

unchanged (no vehicle)

Concentration / amount:

0.4 mL

Day(s)/duration:

6 hours

Adequacy of induction:

highest technically applicable concentration used

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

other: mineral oil

Concentration / amount:

0.4 mL of a 50% w/w mixture

Day(s)/duration:

6 hours

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

- Preliminary irritation study: 4
- Test group: 20
- Control group: 10

Details on study design:

RANGE FINDING TESTS:
A group of 4 animals was used to determine the highest non-irritating concentration (HNIC) of the test substance. The fur was removed by clipping the dorsal area and flanks of each animal. This area was divided in four test sites (two sites on each side of the midline) on each animal. The test substance was applied neat (100%) and also diluted with mineral oil to yield w/w concentrations of 75%, 50% and 25%. Each concentration was applied (0.4 mL) to a test site using an occlusive 25 mm Hilltop Chamber. The sites were wrapped with non-allergic Durapore adhesive tape. After 6 hours of exposure, the chambers were removed and the test sites were cleansed of any residual test substance. Approximately 24 hours after application, each site was evaluated for local reactions (erythema). From these results, the HNIC selected for the challenge phase was a 50% w/w mixture in mineral oil.

MAIN STUDY
A. PREPARATION AND SELECTION OF ANIMALS
Prior to initiation, the fur of a group of animals was removed by clipping the dorsal area and flanks. Animals were weighed and the skin checked to make sure no pre-existing skin irritation was present.

B. INDUCTION EXPOSURE
3 times each week for 3 weeks, 0.4 mL of the undiluted test substance was applied to the left side of each animal using an occlusive 25 mm Hilltop Chamber. The chambers were secured in place and wrapped with non-allergic Durapore adhesive tape to avoid dislocation of the chambers and to minimize loss of the test substance. After an exposure period of 6 hours, the chambers were removed and the test sites were cleansed of any residual test substance. Approximately 24 and 48 hours after each induction application, readings were made of local reactions (erythema).

C. CHALLENGE EXPOSURE
28 days after the first induction dose, 0.4 mL of a 50% w/w mixture of the test substance in mineral oil (HNIC) was applied to a naïve site on the right side of each animals as a challenge dose, under the same occlusive conditions as the induction exposure. Following the 6 hour exposure period, the chambers were removed and the test site were cleansed of residual test substance. These sites were evaluated for sensitization response at approximately 24 and 48 hours after the challenged.

Challenge controls:

10 animals were maintained under identical environmental conditions and treated with the HNIC of the test substance only.

Positive control substance(s):

yes

Remarks:

75% w/w alpha-Hexylcinnamaldehyde in mineral oil

Results and discussion

Positive control results:

4 of 9 positive control animals exhibited signs of sensitization response 24 and 48 hours after challenge. Very faint erythema (0.5) was noted for 4 other sites after challenge.

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

50% w/w

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

very faint erythema (0.5) was observed in 5 animals

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

50% w/w

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

Irriation persisted in one animal

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

50% w/w

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

very faint erythema (0.5) was noted in 6 animals

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

50% w/w

No. with + reactions:

0

Total no. in group:

10

Applicant's summary and conclusion

Interpretation of results:

other: Criteria not met

Remarks:

according to EU CLP (1272/2008 and its amendments)

Conclusions:

In a Buehler test performed in accordance with OECD 406 and according to GLP principles, the substance is not considered a skin sensitizer.

Executive summary:

In a GLP compliant study, in accordance with OECD guideline 406, the skin sensitizing potential of the test substance was evaluated in a Buehler test. The undiluted test substance was topically applied to 20 Hartley Guinea pigs, 2 times each week for a 3 week induction period. 28 days after the first induction dose, a challenge of the test substance at the highest non-irritating concentration (50% w/w in mineral oil) was applied to a naïve site on each animals. A naïve control groups (10 animals) received the challenge alone. Approximately 24 and 48 hours after each induction and challenge dose, the animals were scored for erythema. No sensitizing effects were observed in both the test animals and controls animals. The validity of these results was confirmed by a historical positive control validation study with alpha-Hexylcinnamaldehyde 75% w/w in mineral oil. Based on the results of this study the test substance is not considered to be a skin sensitizer.