Registration Dossier

Administrative data

Description of key information

Skin sensitisation:

Several skin sensitisation tests in guinea pigs are available on methyl ionone from the litterature.

All tests (one GPMT, one FCAT, one Draize, 4 OET and one Photosensitisation test) were negative.

Nevertheless, there is an LLNA on the structural analogue and major contituent of methyl ionone (alpha-isomethyl ionone CAS 127-51-5) that showed some skin sensitising potential, with an EC3 of 21.8%.

Based on this data and the possible read-across from alpha-isomethyl ionone to methyl ionone (see read-across justification document), the fragrance industry chose to adopt a conservative approach and

as a precaution considers methyl ionone as skin sensitizer Cat. 1B as well.

Respiratory sensitisation:

No data were available.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented publication/ study report which meets basic scientific principles
Reason / purpose:
reference to same study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
GPMT
Principles of method if other than guideline:
Guinea pig maximization test according to:
B. Magnusson and A.M. Kligman, The identification of contact allergend by animal assay. The guinea pig miximization-test, ibid., 52, 268-7 (1969).
B. Magnusson and A.M. Kligman, Alelrgic contact dermatitis in the guinea pig, Charles C. Thomas, Springfield, Illinois, 197.
GLP compliance:
not specified
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Available study
Specific details on test material used for the study:
none provided in the publication
Species:
guinea pig
Strain:
Himalayan
Remarks:
white-spotted
Sex:
male/female
Details on test animals and environmental conditions:
weight: 400 - 500 g
Route:
intradermal
Vehicle:
petrolatum
Concentration / amount:
2 injections of 5%
2 injections of 5% in FCA
2 injections of FCA alone
Day(s)/duration:
Once on Day 0
Adequacy of induction:
not specified
Route:
epicutaneous, occlusive
Vehicle:
petrolatum
Concentration / amount:
25% solution in petrolatum of 250mg compound
Day(s)/duration:
Once on day 8 for 48hrs under occlusion
Adequacy of induction:
other: concentration which always causes mild to moderate skin irritation under occlusion
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
petrolatum
Concentration / amount:
specified as sub-irritant
Day(s)/duration:
Once on day 21 for 24hrs
No. of animals per dose:
no data
Details on study design:
Intradermal inductions consisted of 2 injections of 0.1 ml of 5 % test material; 2 injections of 0.1 ml of a 5 % emulsion of test material in Freund's complete adjuvant (FCA); 2 injections of FCA alone on day 0. In addition, 250 mg test material dissolved in petrolatum at 25 % concentration was applied on day 8 under occlusion for 48 hours to a clipped area of the neck.


MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 injections at day 0
- Site: intreadermal
- Concentrations: 5 % in 0.1 ml; with and without Freund's Complete Adjuvant (FCA)
- Control: FCA alone
- in additon: after 8 days 250 mg test material dissolved in petrolatum at 25 % concentration was applied on day 8 under occlusion for 48 hours to a clipped area of the neck.


B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) after study begin: 21
- Exposure period: 24 h
- Site: flank
- Concentrations: subirritant concentration in petrolatum via closed patch
- Evaluation (hr after challenge): 24, 48 h after patch removal
Positive control substance(s):
no
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
subirritant concentration
No. with + reactions:
0
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
subirritant concentration
No. with + reactions:
0
Remarks on result:
no indication of skin sensitisation

no effects observed

Interpretation of results:
GHS criteria not met
Conclusions:
Methyl Ionone did not show any skin sensitizng potential under this Guina Pig Maximisation test when tested intradermally at 5% and topically at 25%.
Executive summary:

The guinea pig maximization test was conducted using outbred Himalayan white-spotted male and female guinea pigs (400-500 grams). Intradermal inductions consisted of 2 injections of 0.1 ml of 5% test material; 2 injections of 0.1 ml of a 5% emulsion of test material in Freund's complete adjuvant (FCA); 2 injections of FCA alone on day 0. In addition, 250 mg test material dissolved in petrolatum at 25% concentration was applied on day 8 under occlusion for 48 hours to a clipped area of the neck. Challenge on day 21 was via a 24 hour closed patch applied to the flank at subirritant concentration. Reactions were read at 24 and 48 hours after removing the patch.No reactions were observed.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented publication/ study report which meets basic scientific principles
Reason / purpose:
reference to same study
Principles of method if other than guideline:
Doses of 0.05 ml of the undiluted compound mixed with the same volume of FCA were injected intradermally into the neck on days 0, 2, 4, 7 and 9 (total dose of 250 mg). The control animals were similarly treated with 5 x 0.05 ml of FCA alone, All the animals were tested epicutaneously on days 21 and 35 as described in the cross-referenced GPMT (key study).
GLP compliance:
not specified
Type of study:
Freund's complete adjuvant test
Justification for non-LLNA method:
Available study
Specific details on test material used for the study:
not provided in the publication
Species:
guinea pig
Strain:
Himalayan
Sex:
male/female
Details on test animals and environmental conditions:
weight: 400 - 500 g
Route:
intradermal
Vehicle:
other: Freund's Complete Adjuvant (FCA)
Concentration / amount:
% times injections of methyl ionone in FCA (1:1). Total dose of 250 mg
Day(s)/duration:
On days 0, 2, 4, 7 and 9
Adequacy of induction:
not specified
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
petrolatum
Concentration / amount:
specified as sub-irritant
Day(s)/duration:
On day 21
Adequacy of challenge:
not specified
No.:
#2
Route:
epicutaneous, occlusive
Vehicle:
petrolatum
Concentration / amount:
specified as sub-irritant
Day(s)/duration:
On day 35
Adequacy of challenge:
not specified
No. of animals per dose:
not specified
Details on study design:
MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 5
- Control group: FCA only
- Site: neck
- Frequency of applications: day 0, 2, 4, 7, 9
- Concentrations: 50 % in 0.1 ml


B. CHALLENGE EXPOSURE
- No. of exposures: 2
- Day(s) after study begin: 21, 35
- Exposure period: 24 h
- Concentrations: subirritant concentration via closed patch
Positive control substance(s):
not specified
Positive control results:
no data
Key result
Reading:
other: both challenges, 1st and 2nd reading at 24 and 48 hours after occlusive patch removal
Group:
test group
Dose level:
sub-irritant level
No. with + reactions:
0
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation

no effects observed

Interpretation of results:
GHS criteria not met
Conclusions:
Methyl Ionone did not show any skin sensitizng potential under this Freund's Complete Adjuvent Test when tested undiluted with FCA (1:1).
Executive summary:

The Freund's complete adjuvant test (FCAT) was conducted using outbred Himalayan white-spotted male and female guinea

pigs weighing 400-500 grams. Induction was via 5 intradermal injections of 0.1 ml of a 50:50 mixture of test material and FCA into the neck

on days 0, 2, 4, 7 and 9. The control animals were similarly treated with 5 x 0.05 ml of FCA alone. Challenge on days 21 and 35 was via a

24 hour closed patch applied to the flank at subirritant concentration. Control animals also challenged on days 21 and 35. No effects were observed.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented publication/ study report which meets basic scientific principles
Reason / purpose:
reference to same study
Principles of method if other than guideline:
J.H. Draize, Appraisal of the safety of chemicals in foods, drugs and cosmetics, Dermal toxicity, The Assoc. of Food and Drug Officials of the United States, Texas State Dept. of Health, Austin, 1959, Pp. 46-59.
GLP compliance:
not specified
Type of study:
Draize test
Justification for non-LLNA method:
Available study
Specific details on test material used for the study:
not provided in the publication
Species:
guinea pig
Strain:
Himalayan
Sex:
male/female
Details on test animals and environmental conditions:
weight: 400 - 500 g
Route:
intradermal
Vehicle:
other: isotonic saline
Concentration / amount:
0.05 ml of a 0.1% solution in saline
Day(s)/duration:
Day 1
Adequacy of induction:
not specified
Route:
intradermal
Vehicle:
other: isotonic saline
Concentration / amount:
0.1 ml of a 0.1% solution in saline
Day(s)/duration:
On 9 alternate days (total dose = 0.95 mg)
Adequacy of induction:
not specified
No.:
#1
Route:
intradermal
Vehicle:
other: saline
Concentration / amount:
0.05 ml of a 0.1% solution in saline
Day(s)/duration:
On day 35
Adequacy of challenge:
not specified
No.:
#2
Route:
intradermal
Vehicle:
other: saline
Concentration / amount:
0.05 ml of a 0.1% solution in saline
Day(s)/duration:
On day 49
Adequacy of challenge:
not specified
Details on study design:
MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 10
- Test groups: 6 - 8
- Frequency of applications: First day and then on 9 alternate days
- Concentrations: first: 0.1 % in 0.05 ml saline; 9 following: 0.1 ml


B. CHALLENGE EXPOSURE
- No. of exposures: 2
- Day(s) after study begin: 35, 49
- Site: intradermal injection
- Concentrations: 0.1 % in 0.05 ml

OTHER
Evaluation criterion was the mean diameter of the papular reactions
Positive control substance(s):
not specified
Positive control results:
no data
Key result
Reading:
other: 1st and 2nd challenges
Group:
test group
Dose level:
0.1%
No. with + reactions:
0
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation

no effetcs observed

Interpretation of results:
not sensitising
Remarks:
Migrated information
Conclusions:
Methyl Ionone did not show any skin sensitizng potential under this skin sensitizing Draize test in guinea pigs when tested at 0.1% intradermally.
Executive summary:

The Draize test was conducted using male and female outbred Himalayan white-spotted guinea pigs weighing 400-500 grams.

0.05 ml test material in normal saline was injected intradermally on day 0 followed by 0.1 ml injection on 9 alternate days for a total dose of

0.95 mg. Intradermal challenge dose of 0.05 ml test material was given on days 35 and 49. Control animals were also challenged

intradermally on days 35 and 49. The evaluation criterion was the mean diameter of the papular reactions. The induction and challenge

concentration was 0.1%. No reactions were observed.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented publication/ study report which meets basic scientific principles
Reason / purpose:
reference to same study
GLP compliance:
not specified
Type of study:
open epicutaneous test
Justification for non-LLNA method:
Available study
Specific details on test material used for the study:
not provided in the publication
Species:
guinea pig
Strain:
Himalayan
Sex:
male/female
Details on test animals and environmental conditions:
- weight: 400 - 500 g
Route:
epicutaneous, open
Vehicle:
unchanged (no vehicle)
Concentration / amount:
0.1 ml of undiluted test item
Day(s)/duration:
Daily during 21 days
Adequacy of induction:
other: Maximum nonirritant and minimal irritating concentrations
No.:
#1
Route:
epicutaneous, open
Vehicle:
unchanged (no vehicle)
Concentration / amount:
0.025 ml of undiluted test item
Day(s)/duration:
Day 21 on contralateral flank
Adequacy of challenge:
other: minimal irritating concentration (confirm biological activity and avoid false results)
No.:
#2
Route:
epicutaneous, open
Vehicle:
unchanged (no vehicle)
Concentration / amount:
0.025 ml of undiluted test item
Day(s)/duration:
Day 35
Adequacy of challenge:
other: minimal irritating concentration (confirm biological activity and avoid false results)
No. of animals per dose:
6 - 8
Details on study design:
Induction consisted of 21 daily open applications of a 0.1-ml aliquot of the test material to an 8 cm2 area on the clipped flank skin of the animals. The reactions were read after 24 hours.
MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 21 following days
- Exposure period: 24 h
- Test groups: 6 - 8
- Site: clipped flank; always same site
- Frequency of applications: daily
- Concentrations: 100 %, 0.1 ml


B. CHALLENGE EXPOSURE
- No. of exposures: 2
- Day(s) after begin of induction: 21, 35
- Exposure period: 24, 48, 72
- Test groups: 6 - 8
- Control group: 6 - 8 untreated animals
- Site: contralateral flank
- Concentrations: 100 %, 0.025 ml
- Evaluation (hr after challenge): 24, 48, 72 h
Challenge controls:
The challenge was conducted on days 21 and 35 via an open application of a 0.025-ml aliquot of the test material that was applied by pipette to a 2 cm2 area on the contralateral flank of the animals. Challenge reactions were evaluated at 24, 48 and 72 hours. Six to eight untreated controls were also treated on days 21 and 35.
Positive control substance(s):
not specified
Positive control results:
no data
Key result
Reading:
other: 1st and 2nd challenge
Group:
test group
Dose level:
100%
No. with + reactions:
0
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation

no effects observed

Interpretation of results:
GHS criteria not met
Conclusions:
Methyl Ionone did not show any skin sensitizng potential under this Guina Pig Open Epicutaneous Test when tested undiluted.
Executive summary:

The open epicutaneous test (OET) was conducted using 6-8 outbred Himalayan white-spotted male and female guinea pigs per

dose. 6-8 untreated animals were used as controls for each test material. Animals weighed 400 to 500 grams. Induction consisted of 21

daily open applications of a 0.1-ml aliquot of the test material to an 8-cm2 area on the clipped flank skin of the animals. The reactions were

read after 24 hours. The challenge was conducted on days 21 and 35 via an open application of a 0.025-ml aliquot of the test material that

was applied by pipette to a 2-cm2 area on the contralateral flank of the animals. Challenge reactions were evaluated at 24, 48 and 72

hours. Six to eight untreated controls were also treated on days 21 and 35. No reactions were reported.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented publication/ study report which meets basic scientific principles
GLP compliance:
no
Type of study:
open epicutaneous test
Justification for non-LLNA method:
Available study
Specific details on test material used for the study:
not provided in the publication
Species:
guinea pig
Strain:
not specified
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Weight at study initiation: 300 - 450 g
Route:
epicutaneous, open
Vehicle:
other: not specified
Concentration / amount:
0.1 ml of 10% test item
Day(s)/duration:
Daily during 21 days
No.:
#1
Route:
epicutaneous, open
Vehicle:
other: not specified
Concentration / amount:
0.025 ml of 10% test item
Day(s)/duration:
Day 21 on contralateral flank
No.:
#2
Route:
epicutaneous, open
Vehicle:
not specified
Concentration / amount:
0.025 ml of 10% test item
Day(s)/duration:
Day 35
No. of animals per dose:
at least 6 animals per dose
Details on study design:
RANGE FINDING TESTS:
- No. of exposures: 1
- Exposure period: 24 h
- Test groups: 6 animals
- Control group: 10 animals
- Site: flank, clipped, 1.4x1.4 cm
- Volume: 0.025 ml
- Concentrations: 3, 10, 30, 100 %
- Evaluation: reactions are read 24 h after application

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 21x in 21 d or 20x in 28 days
- Test groups: 6 / concentration;
- Control group: 10
- Site: flank, clipped, 8 cm2
- Frequency of applications:daily or daily except weekends
- Volume: 0.1 ml
- Concentrations: undiluted
- Evaluation: reactions are read 24 h after application, or end of week

B. CHALLENGE EXPOSURE
- No. of exposures: 2
- Day(s) of challenge: day 21 and 35 after beginning induction
- Test groups: 6 / concentration
- Control group: 10 untreated or pretreated with vehicle (0.1 ml) for 21 days
- Site: contralateral to induction site; 1.4x1.4 cm
- Concentrations: not specified
- Evaluation (hr after challenge): 24 h, 48 h
Positive control substance(s):
not specified
Key result
Reading:
other: both challenges
Group:
test group
Dose level:
10%
No. with + reactions:
0
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation
Interpretation of results:
GHS criteria not met
Conclusions:
Methyl Ionone did not show any skin sensitizng potential under this Guina Pig Open Epicutaneous Test, when tested at 10 %.
Executive summary:

A guinea pig open epicutaneous test (OET) was conducted on groups of 6 - 8 male and female guinea pigs weighting 300 - 450

grams. Daily applications were made for 3 weeks to a clipped 8-cm2 area on the flank of each guinea pig. The test sites were not covered

and the reactions were read 24 hours after each application. A total of 21 applications of 0.1 ml test material in an unspecified vehicle were

made for 21 days. The 10 controls were either left untreated or treated with 0.1 ml of the vehicle for 21 days. At the challenge phase, both

the test and control animals were treated on days 21 and 35 on the contralateral flank with the test material at the minimal irritating

concentration and some lower primary non-irritating concentrations. No effects observed at 10%.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented publication/ study report which meets basic scientific principles
GLP compliance:
not specified
Type of study:
open epicutaneous test
Justification for non-LLNA method:
Available study
Specific details on test material used for the study:
Not provided in the publication
Species:
guinea pig
Strain:
Himalayan
Sex:
male/female
Details on test animals and environmental conditions:
- weight: 400 - 500 g
Route:
epicutaneous, open
Concentration / amount:
10 %
Route:
epicutaneous, open
Concentration / amount:
10 %
No. of animals per dose:
6 - 8
Details on study design:
Induction consisted of 21 daily open applications of a 0.1-ml aliquot of the test material to an 8 cm2 area on the clipped flank skin of the animals. The reactions were read after 24 hours.
MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 21 following days
- Exposure period: 24 h
- Test groups: 6 - 8
- Control group:
- Site: clipped flank; always same site
- Frequency of applications: daily
- Concentrations: 100 %, 0.1 ml


B. CHALLENGE EXPOSURE
- No. of exposures: 2
- Day(s) after begin of induction: 21, 35
- Exposure period: 24, 48, 72
- Test groups: 6 - 8
- Control group: 6 - 8 untreated animals
- Site: contralateral flank
- Concentrations: 10 %, 0.025 ml
- Evaluation (hr after challenge): 24, 48, 72 h
Challenge controls:
The challenge was conducted on days 21 and 35 via an open application of a 0.025-ml aliquot of the test material that was applied by pipette to a 2 cm2 area on the contralateral flank of the animals. Challenge reactions were evaluated at 24, 48 and 72 hours. Six to eight untreated controls were also treated on days 21 and 35.
Key result
Reading:
other: both challenges
Group:
test group
Dose level:
10%
No. with + reactions:
0
Clinical observations:
none
Remarks on result:
no indication of skin sensitisation

no effects observed

Interpretation of results:
GHS criteria not met
Conclusions:
Methyl Ionone did not show any skin sensitizng potential under this Guina Pig Open Epicutaneous Test, when tested at 10%.
Executive summary:

An open epicutaneous test was conducted in guinea pigs. Induction consisted of 21 daily open applications to the shaved flank

of 6-8 guinea pigs per group. One to six experimental and one control group was used. Open challenge applications were made on days

21 and 35. Reactions were read at 24, 48 and 72 hours. No effects observed at 10%.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
other: Original reference not translated (japanese)
Type of study:
guinea pig maximisation test
Species:
guinea pig
Route:
other: no data (report in japanese)
Vehicle:
no data
Concentration / amount:
10%
Day(s)/duration:
no data
Adequacy of induction:
other: no data
No.:
#1
Route:
other: No data
Vehicle:
no data
Concentration / amount:
10%
Day(s)/duration:
no data
Adequacy of challenge:
other: no data
Details on study design:
MAIN STUDY
A. INDUCTION EXPOSURE
- Concentrations: 10 %


B. CHALLENGE EXPOSURE
- Concentrations: 10 %
Key result
Reading:
1st reading
Group:
test group
Dose level:
10%
No. with + reactions:
0

no effects observed

Interpretation of results:
GHS criteria not met
Conclusions:
Methyl ionone was found to be non sensitizer in this Magnusson and Kligman guinea pig maximization test that was conducted at 10% in an unspecified vehicle.
Executive summary:

A Magnusson and Kligman guinea pig maximization test was conducted, and methyl ionone at 10% in an unspecified

vehicle was used for the topical induction and challenge phase. No reactions were reported, and methyl ionone was assigned a score of 0.

No further details were provided

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
April 3, 1980
Reliability:
2 (reliable with restrictions)
Qualifier:
equivalent or similar to
Guideline:
other: Open Epicutaneous Test
Version / remarks:
Similar protocol as for the OET, except for the addition of some irradiation with UV light.
Only the non UV-irradiated part of the photoallergy test is considered here.
Deviations:
yes
Remarks:
UV irradiation for some part, the results of the control area that was not irradiated only is taken into account.
GLP compliance:
not specified
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Available study on a Givaudan quality of Methyl ionone.
Species:
guinea pig
Strain:
not specified
Sex:
male/female
Route:
epicutaneous, open
Vehicle:
not specified
Concentration / amount:
0.1 ml of the test material at 10% is applied to 8 cm 2 of the test area and further irradiated with UV light.
Day(s)/duration:
18 days
Adequacy of induction:
not specified
No.:
#1
Route:
epicutaneous, open
Vehicle:
not specified
Concentration / amount:
0.025 ml of the test material at 10% is applied to 2 cm 2 of both skin sites, for challenging. one flank is irradiated and the other is left non-irradiated as a point of comparison.
Day(s)/duration:
2 days
Adequacy of challenge:
not specified
No. of animals per dose:
8
Details on study design:
Induction phase:

Guinea pigs of both sexes, divided in groups of 8 animals each, are shaved on the upper dorsal area. About one hour later, 0.1 ml of the test material is applied to 8 cm2 of the test area. After this pretreatment, the animals are exposed to the UV-B irradiation for 15' and thereafter to UV-A irradiation for 4 hours (Westing-house black light tubes) from a distance of 25 cm. The application of the test material and the following irradiation are performed every second day, total nine times in 18 days. Skin reading for evidence of erythema is made 24 hours after each application, according to the following scale:
0 = no erythema
1 = slight erythema
2 = moderate erythema
3 = strong erythema


Challenge phase:

After a rest period of ten days, the guinea pigs are shaved on both flanks and the test area marked. 0.025 ml of the test material is applied to 2 cm2 of both skin sites, for challenging. Then, the left flank only is irradiated in the same way as described for induction. Skin reading for evidenc of erythema is made 24 hours and 48 hours after challenging, The reactions on the left and right flank are compared for estimation of the photosensitizing capacity. Moreover, a control group of six animals is tested and read just as the experimental group during the challenge phase.
Challenge controls:
A control group of six animals is tested and read just as the experimental group during the challenge phase.
Positive control substance(s):
not specified
Positive control results:
not specified
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
0.025 mL
No. with + reactions:
0
Total no. in group:
0
Clinical observations:
No erythema
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
0.025 mL
No. with + reactions:
0
Total no. in group:
0
Clinical observations:
No Erythema
Remarks on result:
no indication of skin sensitisation

During both readings (24 and 48 hours) all 8 animals were rated 0 (=no erythema).

No readings were performed during the induction phase since the skin sites used had been stripped.

Interpretation of results:
GHS criteria not met
Conclusions:
Under test conditions used the 10% concentration of RALDEINES (=ISORALDEINE 70) showed no photosensitizing capacity.
Executive summary:

Sensitization was assessed during an associated photosensitization study. Guinea pigs of both sexes, divided in groups of 8 animals each, were shaved on the upper dorsal area. About one hour later, 0.1 ml of 10.0% test material in an undefined vehicle was applied to 8 cm2 of the test area. After the pretreatment, the animals were exposed to UV B-irradiation for 15' and thereafter to UV A-irradiation for 4 hours (Westinghouse black light tubes) from a distance of 25 cm. The application of the test material and the following irradiation were performed every second day, total nine times in 18 days. Skin readings for evidence of erythema were made 24 hours after each application, according to the following scale: 0 = no erythema, 1 = slight erythema, 2 = moderate erythema, 3 = strong erythema. No reading was performed when the skin site used for induction had been stripped. After a rest period of 10 days, the guinea pigs were shaved on both flanks and the test area marked. 0.025 ml of the test material at 10% in an undefined vehicle was applied to 2 cm2 of both skin sites, for challenging. The left flank only was irradiated in the same way as described for induction. Skin reading for evidence of erythema was made 24 and 48 hours after challenging. The reactions on the left and right flank were compared for estimation of the photosensitizing capacity. A control group of 6 animals was tested and read just as the experimental group during the challenge phase.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)
Additional information:

Animal information

In a study to evaluate the skin sensitization potential of methylionone tests like the guinea pig maximization test (GPMT), open epicutanous test (OET), the Draize-Test and Freund’s Complete Adjuvant Test (FCAT) with guinea pigs were compared (Klecak, 1977). For the GPMT, male and female outbred Himalayan guinea pigs were injected intradermally two times with 0.1 ml of 5% methylionone with and without Freund's complete adjuvant (FCA) on day 0. In addition, 250 mg test material dissolved in petrolatum at 25% concentration was applied on day 8 under occlusion for 48 hours to a clipped area of the neck. The Challenge on day 21 was via a 24 hour closed patch at subirritant concentration.  The reading at 24 and 48 hours after removing the patch revealed no sensitizing effects.

The OET was conducted using 6-8 male and female outbred Himalayan guinea pigs while the same number of untreated animals was used as controls (Klecak, 1977). The induction was via 21 daily open applications of 0.1 ml of 100% methylionone to an 8 cm2 area on the clipped flank skin of the animals. The challenge was conducted on days 21 and 35 via open application of 25 µl of a concentration of 100% methylionone to the contralateral flank of the animals. Evaluation of the sites at 24, 48 and 72 h after challenge revealed no sensitizing effects.
The Draize test was conducted using male and female outbred Himalayan guinea pigs (Klecak, 1977). 0.05 ml of 0.1% methylionone in normal saline was injected intradermally on day 0 followed by 0.1 ml injection on 9 alternate days for a total dose of 0.95 mg. A following intradermal challenge dose of 0.05 ml was given on days 35 and 49. The evaluation criterion was the mean diameter of the papular reactions. As a result, no sensitization to skin was detected.

The Freund's complete adjuvant test (FCAT) was conducted using male and female outbred Himalayan guinea pigs (Klecak, 1977). The induction was performed via 5 intradermal injection of 0.1 ml of a 50:50 mixture of methylionone and FCA into the neck of the animals on days 0, 2, 4, 7 and 9. The control animals were similarly treated with 5 x 0.05 ml of FCA alone. The challenge was via a 24 hour closed patch applied to the flank of test and control animals at subirritant concentration on days 21 and 35. No sensitizing effect was detected.

In another study by the same author (Klecak, 1985), the basic scope and limits of the FCAT and the OET were comparatively discussed. Within a table, the author summarized the predictive value of the OET compared to human tests such as the Human Maximization Test (HMT) and the Repeat Insult Patch Test (RIPT) for a total of 290 substances including methyl ionone.

For methyl ionone, the result of the OET testing with guinea pig was compared to that of the HMT reported by Kligmann AM (1966). The OET was conducted with 6 male and femal guinea pigs which received 21 dermal applications of 0.1 ml methylionone on an area of 8 cm2 on the flank in 21 days or 20 applications in 28 days. The challenge consisted of two applications on days 21 and 35 and evaluation was done 24 and 48 later. No sensitizing effects were detected.

In another study of the same author (Klecak, 1979), also an OET was conducted using 6-8 male and female outbred Himalayan guinea pigs while the same number of untreated animals was used as controls. Induction consisted of 21 daily open applications of a 0.1-ml aliquot of the test material to an 8 cm2 area on the clipped flank skin of the animals. The reactions were read after 24 hours. The challenge was conducted on days 21 and 35 via open application of 25 µl of a concentration of 10% methylionone to the contralateral flank of the animals. Evaluation of the sites at 24, 48 and 72 h after challenge revealed no sensitizing effects.

In a third study of the same author (Klecak, 1985), also an OET was conducted on groups of 6 - 8 male and female guinea pigs weighting 300 - 450 grams. Daily applications were made for 3 weeks to a clipped 8-cm2 area on the flank of each guinea pig. The test sites were not covered and the reactions were read 24 hours after each application. A total of 21 applications of 0.1 ml test material in an unspecified vehicle were made for 21 days. The 10 controls were either left untreated or treated with 0.1 ml of the vehicle for 21 days. At the challenge phase, both the test and control animals were treated on days 21 and 35 on the contralateral flank with the test material at the minimal irritating concentration and some lower primary non-irritating concentrations. No effects observed at 10%.


Another GPMT was described in a Japanese publication and used concentrations of 10% of methylionone for induction and challenge. As result, no sensitizing effects were reported (Ishihara. 1986).

Finally, sensitization of methyl ionone was also assessed during an associated photosensitization study (Givaudan, 1098). Guinea pigs of both sexes, divided in groups of 8 animals each, were shaved on the upper dorsal area. About one hour later, 0.1 ml of 10.0% test material in an undefined vehicle was applied to 8 cm2 of the test area. After the pretreatment, the animals were exposed to UV B-irradiation for 15' and thereafter to UV A-irradiation for 4 hours (Westinghouse black light tubes) from a distance of 25 cm. The application of the test material and the following irradiation were performed every second day, total nine times in 18 days. Skin readings for evidence of erythema were made 24 hours after each application, according to the following scale: 0 = no erythema, 1 = slight erythema, 2 = moderate erythema, 3 = strong erythema. No reading was performed when the skin site used for induction had been stripped. After a rest period of 10 days, the guinea pigs were shaved on both flanks and the test area marked. 0.025 ml of the test material at 10% in an undefined vehicle was applied to 2 cm2 of both skin sites, for challenging. The left flank only was irradiated in the same way as described for induction. Skin reading for evidence of erythema was made 24 and 48 hours after challenging. The reactions on the left and right flank were compared for estimation of the photosensitizing capacity. A control group of 6 animals was tested and read just as the experimental group during the challenge phase. No reaction and no erythema was observed with or without UV exposure.

Nevertheless, there is an LLNA on the structural analogue and major contituent of methyl ionone (alpha-isomethyl ionone CAS 127-51-5) that showed some skin sensitising potential.

The Local Lymph Node Assay (Betts, 2005) was conducted in female mice. Each animal received a daily topical application of 2.5%, 5%, 10%, 25% or 50% (w/v) alpha-iso-methylionone in EtOH : DEP (3:1) on the dorsal surface of each ear for 3 consecutive days. Three days after the third topical application all mice were injected intravenously through the tail vein with 250 μl sterile saline (PBS) containing 20 μCi 3Hmethylthymidine (3H-thymidine). All mice were sacrificed five hours after the intravenous injection. Draining auricular lymph nodes were excised and were pooled for each experimental group. 3H-TdR incorporation was then measured by beta-scintillation counting and stimulation indices were determined for each experimental group. alpha-iso-Methylionone was considered to be a potential sensitizer under the conditions of the test with an EC3 value of 21.8% (5450 μg/cm2).

Based on this data and the possible read-across from alpha-isomethyl ionone to methyl ionone (see read-across justification document), the fragrance industry chose to adopt a conservative approach and as a precaution considers methyl ionone as skin sensitizer Cat. 1B as well.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The present data on skin sensitisation do not fulfill the criteria laid down in 67/548/EEC and 1272/2008/EEC.

Nevertheless, there is data on the structural analogue and also major constituent alpha-isomethal ionone, that indicate a certain skin sensitising potential (Betts, 2005). Based on this data and the possible read-across from alpha-isomethyl ionone to methyl ionone (see read-across justification document), the fragrance industry chose to adopt a conservative approach and classify as a precaution methyl ionone (Skin Sens. 1B) as well.