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Administrative data

Description of key information

Guinea pig. Not sensitising. OECD 406; Reliability = 1

Guinea pig. Not sensitising. OECD 406; Reliability = 1

Guinea pig. sensitising. OECD 406; Reliability = 1

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:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
other: JMAFF 12-Nousan-8147 (2000)
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
LLNA study was not performed since in vivo guinea pig data were already available for the test substance.
Species:
guinea pig
Strain:
other: Hartley albino
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Elm Hill Breeding Labs, Chelmsford, MA
- Age at study initiation: Preliminary Irritation Group: Young adult (5 weeks), Test and Test Vehicle Control Groups: Young adult (4 weeks)
- Weight at study initiation: 373-439 grams
- Housing: The animals were gang housed in plastic solid bottom polycarbonate cages or stainless steel solid bottom cages
- Diet: Approximately 20 grams per day
- Water: ad libitum
- Acclimation period: 14 or 19 days

ENVIRONMENTAL CONDITIONS
- Temperature: 19-23°C
- Humidity: 53-65%
- Air changes (per hr): 12
- Photoperiod (hrs dark / hrs light): 12
Route:
intradermal
Vehicle:
other: 2% w/v solution of CMC in distilled water
Concentration / amount:
The test animals received six intradermal injections (0.1 mL each) in the shaved suprascapular region as follows:
Emulsion of Freund’s Adjuvant Complete (50% v/v in distilled water), 1% w/w mixture of test substance in a 2% w/v solution of CMC in distilled water, 1% w/w mixture of test substance in an emulsion of Freund’s Adjuvant Complete (50% v/v in distilled water)
Day(s)/duration:
24 and 48 hours
Route:
other: Topical
Vehicle:
other: 2% w/v solution of CMC in distilled water
Concentration / amount:
0.5 grams of 65% w/w mixture of the test substance in a 2% w/v solution of CMC in distilled water
Day(s)/duration:
48 hours
Route:
other: Topical
Vehicle:
other: 2% w/v solution of CMC in distilled water
Concentration / amount:
0.5 grams of a 65% w/w mixture of the test substance in a 2% w/v solution of CMC in distilled water (HNIC), 0.5 mL of a 22% w/w mixture in a 2% w/v solution of CMC in distilled water and 100% of a 2% w/v solution of CMC in distilled water
Day(s)/duration:
24 hours
No. of animals per dose:
Preliminary Irritation Testing: 12
Test Group: 20
Test Vehicle Control Group: 10
Details on study design:
RANGE FINDING TESTS:
Preliminary irritation testing
Preliminary Intradermal Injection: Prior to the induction phase, a group of four animals was used to determine the concentration of the test substance which produced faint to moderate irritation via intradermal injection. The fur was removed by clipping the suprascapular region of each guinea pig. This area was divided into six test sites (three sites on each side of the midline) on each animal. Each guinea pig received six intradermal injections (0.1 mL each); three concentrations (1, 3, and 5%) of the test substance in a 2% w/v solution of CMC in distilled water and the same concentrations in an emulsion of Freund’s Adjuvant Complete. All preparations in Freund’s Adjuvant Complete were thoroughly mixed with a tissue homogenizer prior to application. Approximately 24 and 48 hours after the injections, each site was evaluated for local reactions (erythema).
Preliminary Topical: Prior to the topical induction, a group of four animals was used to determine the irritation potential of the test substance to be used during the topical induction. The previously clipped flank area of each guinea pig was divided into two sites (one site on each side of the midline). The test substance was applied mixed with a 2% w/v solution of CMC in distilled water to yield w/w concentrations of 65%1 and 49%. Each concentration (0.5 g or mL) was applied to a 2 cm x 4 cm, 2-ply gauze patch and placed on one of the two test sites. The patch was covered with plastic wrap and secured in place with non-allergenic Durapore™ adhesive tape to avoid dislocation of the patch and to minimize loss of the test substance. After a 48 hour exposure period, the patches were removed and the test sites were gently cleansed of any residual test substance. One hour after patch removal readings were made of local reactions (erythema) according to the scoring system.
Highest Non-irritating Concentration (HNIC): Prior to the challenge phase, a group of four animals was used to determine the highest non-irritating concentration. The fur was removed by clipping the flanks of each guinea pig. The test substance was mixed with a 2% w/v solution of CMC in distilled water to yield w/w concentrations of 65%, 49%, 33% and 16%. Each concentration (0.5 g or mL) was applied to an occlusive 25 mm Hill Top Chamber and applied to the appropriate test site. The sites were wrapped with non-allergenic Durapore™ adhesive tape. After 24 hours of exposure, the chambers were removed and the test sites were gently cleansed of any residual test substance. Approximately 24 and 48 hours after patch removal, each site was evaluated for local reactions (erythema) according to the scoring system.
Based on these findings, the concentration which produced faint to moderate irritation (1-2) selected for the intradermal induction was a 1%1 w/w mixture in a 2% w/v solution of CMC in distilled water. That which produced no irritation selected for the topical induction was a 65% w/w mixture in a 2% w/v solution of CMC in distilled water. Due to the lack of irritation produced during preliminary topical induction testing, a pretreatment of sodium lauryl sulfate was applied to all animals prior to the topical induction test substance application. The HNIC (the highest concentration that produced responses in four guinea pigs no more severe than two scores of 0.5 and two scores of zero) selected for the challenge phase was a 65% w/w mixture in a 2% w/v solution of CMC in distilled water.
MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 6
- Exposure period: 24 and 48 hours for Intradermal induction and 48 hours for topical induction
- Test groups: 1
- Control group: 1
- Site: Suprascapular region
- Concentrations: Induction phase: Emulsion of Freund’s Adjuvant Complete (50% v/v in distilled water), 1% w/w mixture of test substance in a 2% w/v solution of CMC in distilled water, 1% w/w mixture of test substance in an emulsion of Freund’s Adjuvant Complete (50% v/v in distilled water)
-Frequency of application: once in intradermal and once in topical
-Duration: 7 days

B. CHALLENGE EXPOSURE
- No. of exposures: 6
- Day of challenge: 21
- Exposure period: 24 hours
- Test groups: 1
- Control group: 1
- Site: Right and left flank of each animal
- Concentrations: 0.5 grams of a 65% w/w mixture of the test substance in a 2% w/v solution of CMC in distilled water (HNIC), 0.5 mL of a 22% w/w mixture in a 2% w/v solution of CMC in distilled water and 0.5 mL of a 2% w/v solution of CMC in distilled water
- Evaluation (hr after challenge): 24 and 48 hours
Challenge controls:
2% w/v solution of CMC in distilled water
Positive control substance(s):
yes
Remarks:
Alpha-Hexylcinnamaldehyde
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
65% w/w in a 2% w/v solution of CMC in distilled water
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted for four of twenty test sites 24 hours after challenge.
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
65% w/w in a 2% w/v solution of CMC in distilled water
No. with + reactions:
0
Total no. in group:
20
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
22% w/w in a 2% w/v solution of CMC in distilled water
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted for three of twenty test sites 24 hours after challenge.
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
22% w/w in a 2% w/v solution of CMC in distilled water
No. with + reactions:
0
Total no. in group:
20
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100% of a 2% w/v solution of CMC in distilled water
No. with + reactions:
0
Total no. in group:
20
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
100% of a 2% w/v solution of CMC in distilled water
No. with + reactions:
0
Total no. in group:
20
Reading:
1st reading
Group:
positive control
Remarks on result:
other: Historical positive control data were used. Positive indication of sensitization response was observed.
Reading:
1st reading
Group:
negative control
Dose level:
control group for 65 % w/w
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Group:
negative control
Dose level:
control group for 22% w/w
Remarks on result:
no indication of skin sensitisation

Table 1: Incidence of the sensitization response noted after challenge

Dose

Incidence with Skin Reactions1

Test animals

Test Vehicle Control/ Naïve Animals

Hours

24

48

24

48

65% w/w in a

2% w/v solution of CMC in distilled water

0/20

0/20

0/10

0/10

22% w/w in a

2% w/v solution of CMC in distilled water

0/20

0/20

0/10

0/10

100% of a 2% w/v solution of CMC in distilled water

0/20

0/20

0/10

0/10

1 Animals with scores greater than 0.5

Table 2: Severity of the sensitization response noted after challenge

Dose

Severity2

Test animals

Test Vehicle Control/ Naïve Animals

Hours

24

48

24

48

65% w/w in a

2% w/v solution of CMC in distilled water

0.10

0.00

0.00

0.00

22% w/w in a

2% w/v solution of CMC in distilled water

0.08

0.00

0.10

0.00

100% of a 2% w/v solution of CMC in distilled water

0.00

0.00

0.00

0.00

2 The test substance, as received, was a solid. To enhance skin contact, the test substance was moistened with a 2% w/v solution of CMC in distilled water prior to application.

Interpretation of results:
GHS criteria not met
Conclusions:
Non sensitizer
Executive summary:

The study was conducted according to guidelines, U.S. EPA OPPTS 870.2600 and OECD Guideline 406. A Magnusson-Kligman maximization test was conducted with guinea pigs to determine the potential for test substance to invoke dermal skin sensitization reactions. The study was conducted using four stages; preliminary irritation screens, a two-stage induction phase, and a challenge phase. Preliminary irritation testing was performed on 12 animals to determine appropriate concentrations of the test substance that could be used for both intradermal and topical induction as well as topical challenge.

An emulsion of 50% v/v Freund’s Adjuvant Complete in distilled water was used during the intradermal injection screening and the injection induction phases. This emulsion was thoroughly mixed using a stir plate and is referred to throughout the report as an emulsion of Freund’s Adjuvant Complete.

The first induction phase involved six intradermal injections into the suprascapular area of each of 20 guinea pigs. These doses were comprised of pairs of injections of the test substance in a 2% w/v solution of carboxymethylcellulose (CMC) in distilled water (1% w/w), the test substance (1% w/w) combined with an emulsion of Freund’s Adjuvant Complete, as well as an emulsion of Freund’s Adjuvant Complete alone. A sham control group (ten animals) was maintained under the same environmental conditions and received injections of a 2% w/v solution of CMC in distilled water, a 2% w/v solution of CMC in distilled water (50% w/w) combined with an emulsion of Freund’s Adjuvant Complete, as well as an emulsion of Freund’s Adjuvant Complete alone. Approximately 24 and 48 hours after the injections, all sites were evaluated for an irritation response (erythema).

Approximately one week later, the second phase of induction was conducted. Due to a lack of irritation produced during preliminary testing, all animals received a pretreatment of sodium lauryl sulfate prior to test substance application. A 65% w/w mixture of the test substance in a 2% w/v solution of CMC in distilled water (test group) or a 2% w/v solution of CMC in distilled water (test vehicle control group) was then applied topically for a period of 48 hours to the area encompassing the intradermal injection sites. Approximately one hour after the topical induction patches were removed, all animals were scored for erythema.

Approximately two weeks later, a primary challenge consisting of three occluded applications was conducted on each animal. One Hill Top Chamber containing 0.5 mL of a 2% w/v solution of CMC in distilled water was applied to a naive site on the right middle flank of each animal. The remaining two Hill Top Chambers containing 0.5 mL of the HNIC (Highest Non-Irritating Concentration, determined in the preliminary irritation screen to be a 65% w/w mixture in a 2% w/v solution of CMC in distilled water) of the test substance and 0.5 mL of a 33% dilution of the HNIC (22% w/w mixture in a 2% w/v solution of CMC in distilled water) were positioned on two naive sites on the left front and rear flank, respectively, for approximately 24 hours. The test vehicle control group was also treated with the test substance and test vehicle at challenge. Approximately 24 and 48 hours after challenge patch removal, all animals were scored for a sensitization response (erythema).

The results showed no incidence of skin reactions at 24 and 48 hours after challenge doses of 65% w/w in a 2% w/v solution of CMC in distilled water, 22% w/w in a 2% w/v solution of CMC in distilled water and 100% of a 2% w/v solution of CMC in distilled water. Based on these results, test substance is considered not to be a contact skin sensitizer.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
other: JMAFF 12-Nousan-8147 (2000)
Deviations:
no
Qualifier:
according to
Guideline:
other: Official Journal of the European Communities. Methods for the Determination of Toxicity and Health Effects, Part B.6 (Skin Sensitization) Commission Regulation (EC) No. 440/2008
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
LLNA was not performed since in vivo guinea pig data was already available for the test substance.
Species:
guinea pig
Strain:
other: Hartley albino
Sex:
male
Route:
intradermal
Vehicle:
polyethylene glycol
Concentration / amount:
The test animals received six intradermal injections (0.1 mL each) in the shaved suprascapular region as follows: Emulsion of Freund’s Adjuvant Complete (50% v/v in distilled water), 1% w/w mixture of test substance in polyethylene glycol, 1% w/w mixture of test substance in an emulsion of Freund’s Adjuvant Complete (50% v/v in distilled water)
Day(s)/duration:
24 and 48 hours
Route:
other: Topical
Vehicle:
polyethylene glycol
Concentration / amount:
53% w/w mixture of the test substance in polyethylene glycol
Day(s)/duration:
48 hours
Route:
other: Topical
Vehicle:
polyethylene glycol
Concentration / amount:
0.5 mL of a 70% w/w mixture of the test substance in polyethylene glycol and a 23% w/w mixture of the test substance in polyethylene glycol
Day(s)/duration:
24 hours
No. of animals per dose:
Preliminary Irritation Testing: 12
Test Group: 20
Test Vehicle Control Group: 10
Challenge controls:
polyethylene glycol
Positive control substance(s):
yes
Remarks:
alpha-Hexylcinnamaldehyde
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
70% w/w in polyethylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted for four test sites 24 hours after challenge patch removal
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
70% w/w in polyethylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted for one test site 48 hours after challenge patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
23% w/w in polyethylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted for three test sites 24 hours after challenge patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
23% w/w in polyethylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted for one test site 48 hours after challenge patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100% polyethylene glycol):
No. with + reactions:
0
Total no. in group:
20
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
100% polyethylene glycol):
No. with + reactions:
0
Total no. in group:
20
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Group:
positive control
Remarks on result:
other: Historical control positive control data were used. Postive sensitization responses were noted.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
control group for 70% w/w
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Group:
negative control
Dose level:
control group for 23% w/w
Remarks on result:
no indication of skin sensitisation
Interpretation of results:
GHS criteria not met
Conclusions:
Non sensitizer
Executive summary:

The study was conducted according to guidelines, U.S. EPA OPPTS 870.2600 and OECD Guideline 406. A Magnusson-Kligman test was conducted with guinea pigs to determine the potential for test substance to invoke dermal skin sensitization reactions.

The study was conducted using four stages; preliminary irritation screens, a two-stage induction phase, and a challenge phase. Preliminary irritation testing was performed on twelve animals to determine appropriate concentrations of the test substance that could be used for both intradermal and topical induction as well as topical challenge.

The first induction phase involved six intradermal injections into the suprascapular area of each of twenty guinea pigs. These doses were comprised of pairs of injections of the test substance in polyethylene glycol (1% w/w), the test substance (1% w/w) combined with an emulsion of Freund’s Adjuvant Complete, as well as an emulsion of Freund’s Adjuvant Complete alone. A sham control group (ten animals) was maintained under the same environmental conditions and received injections of polyethylene glycol, polyethylene glycol (50% w/w) combined with an emulsion of Freund’s Adjuvant Complete, as well as an emulsion of Freund’s Adjuvant Complete alone. Approximately 24 and 48 hours after the injections, all sites were evaluated for an irritation response (erythema).

Approximately one week later, the second phase of induction was conducted. The test substance as a 53% w/w mixture in polyethylene glycol (test group) or polyethylene glycol (test vehicle control group) was applied topically for a period of 48 hours to the area encompassing the intradermal injection sites. Approximately one hour after the topical induction patches were removed, all animals were scored for erythema. Approximately two weeks later, a primary challenge consisting of three occluded applications was conducted on each animal. One Hill Top Chamber containing 0.5 mL of polyethylene glycol was applied to a naive site on the right middle flank of each animal. The remaining two Hill Top Chambers containing 0.5 mL of the HNIC (Highest Non-Irritating Concentration, determined in the preliminary irritation screen to be a 70% w/w mixture in polyethylene glycol) of the test substance and 0.5 mL of a 33% dilution of the HNIC (23% w/w mixture in polyethylene glycol) were positioned on two naive sites on the left front and rear flank, respectively, for approximately 24 hours. The test vehicle control group was also treated with the test substance and test vehicle at challenge. Approximately 24 and 48 hours after challenge patch removal, all animals were scored for a sensitization response (erythema). The results showed no incidence of skin reactions at 24 and 48 hours in all animals tested at concentration of 70% w/w in polyethylene glycol and 23% w/w in polyethylene glycol. Based on the results of this study, test substance is not considered to be a contact skin sensitizer.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
other: JMAFF 12-Nousan-8147 (2000)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
LLNA study was not performed since in vivo guinea pig data were already available for the test substance.
Species:
guinea pig
Strain:
other: Hartley albino
Sex:
male
Route:
intradermal
Vehicle:
propylene glycol
Concentration / amount:
Concentration / amount
The test animals received six intradermal injections (0.1 mL each) in the shaved suprascapular region as follows: Emulsion of Freund’s Adjuvant Complete (50% v/v in distilled water), 1% w/w mixture of test substance in propylene glycol, 1% w/w mixture of test substance in an emulsion of Freund’s Adjuvant Complete (50% v/v in distilled water)
Day(s)/duration:
24 and 48 hours
Route:
other: Topical
Vehicle:
propylene glycol
Concentration / amount:
Five tenths of a gram of a 65% w/w of the test substance in propylene glycol
Day(s)/duration:
48 hours
Route:
other: Topical
Vehicle:
propylene glycol
Concentration / amount:
0.5 mL of 49% w/w mixture of the test substance in propylene glycol and 0.5 mL of a 16% w/w mixture in propylene glycol
Day(s)/duration:
24 hours
No. of animals per dose:
Preliminary Irritation Testing: 12
Test Group: 20
Test Vehicle Control Group: 10
Rechallenge Vehicle Control Group: 10
Challenge controls:
propylene glycol
Positive control substance(s):
yes
Remarks:
AlphaHexylcinnamaldehyde
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
49% w/w in propylene glycol
No. with + reactions:
6
Total no. in group:
20
Clinical observations:
Very faint to faint erythema (0.5-1) was noted at nineteen of twenty test sites 24 hours after patch removal.
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
49% w/w in propylene glycol
No. with + reactions:
1
Total no. in group:
20
Clinical observations:
Very faint to faint erythema (0.5-1) was noted at nineteen of twenty test sites 48 hours after patch removal. Faint erythema persisted at one test site at 48 hours. Desquamation was noted at five test sites 48 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
16% w/w in propylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted at seven of twenty test sites 24 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
16% w/w in propylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted at seven of twenty test sites 48 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100% propylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted at two of twenty test sites 24 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
100% propylene glycol
No. with + reactions:
0
Total no. in group:
20
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
33% w/w in propylene glycol
No. with + reactions:
6
Total no. in group:
20
Clinical observations:
Very faint to faint erythema (0.5-1) was noted at seventeen of twenty test sites 24 hours after patch removal.
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
test group
Dose level:
33% w/w in propylene glycol
No. with + reactions:
3
Total no. in group:
20
Clinical observations:
Very faint to faint erythema (0.5-1) was noted at seventeen of twenty test sites 48 hours after patch removal. Faint erythema persisted at three test sites at 48 hours. Desquamation was noted at nine test sites 48 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
10% w/w in propylene glycol
No. with + reactions:
1
Total no. in group:
20
Clinical observations:
Very faint to faint erythema (0.5-1) was noted at thirteen of twenty test sites 24 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
test group
Dose level:
10% w/w in propylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint to faint erythema (0.5-1) was noted at thirteen of twenty test sites 48 hours after patch removal. Desquamation was noted at six test sites 48 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
100% propylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted at five of twenty test sites 24 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
test group
Dose level:
100% propylene glycol
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Very faint erythema (0.5) was noted at five of twenty test sites 48 hours after patch removal.
Remarks on result:
no indication of skin sensitisation
Interpretation of results:
Category 1 (skin sensitising) based on GHS criteria
Conclusions:
Contact Skin Sensitizer
Executive summary:

The study was conducted according to guidelines, U.S. EPA OPPTS 870.2600 and OECD Guideline 406. A Magnusson-Kligman test was conducted with guinea pigs to determine the potential for test substance to invoke dermal skin sensitization reactions. The study was conducted using four stages; preliminary irritation screens, a two-stage induction phase, and a challenge phase. Preliminary irritation testing was performed on twelve animals to determine appropriate concentrations of the test substance that could be used for both intradermal and topical induction as well as topical challenge.

An emulsion of 50% v/v Freund’s Adjuvant Complete in distilled water was used during the intradermal injection screening and the injection induction phases. This emulsion was thoroughly mixed using a stir plate and is referred to throughout the report as an emulsion of Freund’s Adjuvant Complete.

The first induction phase involved six intradermal injections into the suprascapular area of each of 20 guinea pigs. These doses were comprised of pairs of injections of the test substance in propylene glycol (1% w/w), the test substance (1% w/w) combined with an emulsion of Freund’s Adjuvant Complete, as well as an emulsion of Freund’s Adjuvant Complete alone. A sham control group (ten animals) was maintained under the same environmental conditions and received injections of propylene glycol, propylene glycol (50% w/w) combined with an emulsion of Freund’s Adjuvant Complete, as well as an emulsion of Freund’s Adjuvant Complete alone. Approximately 24 and 48 hours after the injections, all sites were evaluated for an irritation response (erythema).

Approximately one week later, the second phase of induction was conducted. The test substance as a 65% w/w mixture in propylene glycol (test group) or propylene glycol (test vehicle control group) was applied topically for a period of 48 hours to the area encompassing the intradermal injection sites. Approximately one hour after the topical induction patches were removed, all animals were scored for erythema.

Approximately two weeks later, a primary challenge consisting of three occluded applications was conducted on each animal. One Hill Top Chamber containing 0.5 mL of propylene glycol was applied to a naive site on the right middle flank of each animal. The remaining two Hill Top Chambers containing 0.5 mL of the HNIC (Highest Non-Irritating Concentration, determined in the preliminary irritation screen to be a 49% w/w mixture in propylene glycol) of the test substance and 0.5 mL of a 33% dilution of the HNIC (16% w/w mixture in propylene glycol) were positioned on two naive sites on the left front and rear flank, respectively, for approximately 24 hours. The test vehicle control group was also treated with the test substance and test vehicle at challenge. Approximately 24 and 48 hours after challenge patch removal, all animals were scored for a sensitization response (erythema).

Due to an irritation response observed in both the test and sham control group following the primary challenge with test substance, it was necessary to conduct a rechallenge using a lower concentration of that test substance. Positive reactions (scores of 1) seen in the sham control animals during the challenge phase indicated an irritant response to test substance such that the HNIC had been exceeded. Seven days after the primary challenge, a rechallenge was conducted using the original test animals and a new naive control group (10 animals). The naive control animals used for the rechallenge did not receive intradermal injections and a topical induction application as the original sham control animals had. A 33% w/w mixture of test substance in propylene glycol was selected as the HNIC for rechallenge. New dose sites were selected on each test animal for the re-challenge. A quantity equal to 0.5 mL of propylene glycol was applied to one chamber and positioned on the right middle flank. The remaining two chambers containing 0.5 mL of a 33% w/w mixture of the test substance in propylene glycol (HNIC) and 0.5 mL of a 10% w/w mixture in propylene glycol (33% dilution of the HNIC) were positioned on the left front and rear flank, respectively. The same procedures for application and scoring as described for the primary challenge were used. Approximately 24 and 48 hours after rechallenge application the animals were scored for local reactions (erythema).

The results showed skin reactions with score greater than 0.5 was observed in 6 of 20 animals when challenged for 24 hours with a dose of 49% w/w in propylene glycol and 1 of 20 animals after challenged for 48 hours with a dose of 49% w/w in propylene glycol. Even in rechallenge phase, skin reactions with score greater than 0.5 was observed in 6 of 20 animals with a dose of 33% w/w in propylene glycol rechallenged for 24 hours and 3 of 20 animals after challenged for 48 hours with a dose of 33% w/w in propylene glycol for 48 hours. 1 of 20 animals also developed skin reactions with score greater than 0.5 when rechallenged with a dose of 10% w/w in propylene glycol for 24 hours. These results conclude that the test susbstance is a contact skin sensitizer.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available (further information necessary)
Additional information:

The dermal sensitization potential of the test substance was evaluated in guinea pigs by the Magnusson and Kligman maximisation method according to OECD guideline 406 in male Hartley albino guinea pigs. The percentage of sensitisation at 24 and/or 48 hours for the test article animals challenged with 65% or 22% of test substance was 0%. Under the conditions of this study, the test substance did not possess skin sensitising potential. Tests with another test batch of the test substance also showed no skin sensitising potential.

A third test batch of the test substance (a batch not further developed for registration) was found to have a sensitization potential.

Justification for classification or non-classification

There was no evidence of dermal sensitisation in guinea pigs. The substance does not need to be classified for skin sensitisation according to EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.