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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2005-10-31 - 2005-12-16
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report Date:
2006

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
Type of study:
Buehler test
Justification for non-LLNA method:
Data requirement addressed using existing test data, not specifically tested in order to comply with REACH.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid: viscous
Details on test material:
- Name of test material (as cited in study report): OS 13 199711. IX.5 LT
- Substance type: organic
- Physical state: viscous brown liquid
- Receipt date: 2005-10-24

In vivo test system

Test animals

Species:
guinea pig
Strain:
Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals, Inc., Scottdale, Pennsylvania;
- Age at study initiation: 7 weeks (males), 8 weeks (females);
- Weight at study initiation: 321-403 g (males), 307-377 g (females);
- Housing: individually in suspended stainless steel cages;
- Diet (e.g. ad libitum): ad libitum, PMI Certified Guinea Pig Chow #5026 (PMI Nutrition International);
- Water (e.g. ad libitum): ad libitum (municipal tap water treated by reverse osmosis);
- Acclimation period: 5 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-23
- Humidity (%): 37-68
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES:
2005-11-16 - 2005-12-16

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
other: mineral oil
Concentration / amount:
Test group: 100 % (induction and challenge);
HCA test group: 5 % w/v in ethanol (induction), 2.5 % and 1.0 % w/v in acetone (challenge);
Challenge control group: 100 % (challenge);
HCA challenge control group: 2.5 % and 1.0 % w/v in acetone (challenge).
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
other: mineral oil
Concentration / amount:
Test group: 100 % (induction and challenge);
HCA test group: 5 % w/v in ethanol (induction), 2.5 % and 1.0 % w/v in acetone (challenge);
Challenge control group: 100 % (challenge);
HCA challenge control group: 2.5 % and 1.0 % w/v in acetone (challenge).
No. of animals per dose:
Test group: 10 males, 10 females;
HCA test group: 5 males, 5 females;
Challenge control group: 5 males, 5 females;
HCA challenge control group: 5 males, 5 females.
Details on study design:
RANGE FINDING TESTS:
The results of the range-finding study indicated that a test article concentration of 100% (as received) was considered appropriate for induction and challenge as the 100 % concentration was the highest non-irritating concentration.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 3;
- Exposure period: day 0, 7 and 14;
- Test and control groups: On the day prior to each dose administration, the guinea pigs had the hair removed with a small animal clipper. Care was taken to avoid abrading the skin during the clipping. A 0.3 mL dose of the appropriate test material was placed on a 25 mm Hilltop chamber backed by adhesive tape (occlusive patch). The chambers were then applied to the clipped surface as quickly as possible.
Following chamber application, the trunk of the animal was wrapped with elastic wrap which was secured with adhesive tape to prevent removal of the chamber and the animal was returned to its cage;
- Site: back (left side);
- Frequency of applications: single application on day of induction;
- Duration: 6 hours;
- Concentrations: test article (100 %), positive control substance (5% in ethanol).

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 28
- Exposure period: 6 hours
- Test groups: On the day prior to challenge dose administration, the test, HCA test, challenge control and HCA challenge control animals were weighed and the hair was removed from the right side of the animals. On the day following clipping (day 28), chambers were applied to the right side of the back.
- Control group: yes
- Site: back (right side);
- Concentrations: test article (100 %), positive control substance (2.5 % and 1.0 % in acetone).
- Evaluation (hr after challenge): 24 and 48 hours

OTHER:
Approximately six hours after chamber application, the binding materials were removed. The test sites of the test and challenge control animals were wiped with gauze moistened in mineral oil. USP followed by dry gauze, followed by gauze moistened in deionized water followed by dry gauze. The test sites of the test, HCA test, challenge control and HCA challenge control animals were then wiped with gauze moistened in deionized water, followed by dry gauze, to remove test article residue. The animals were then returned to their cages.
Note: At the time of binding removal, it was noticed that the test article reacted with the tape on the hilltop chamber. As a result, 5 animals had sticky residue outside the test site and the other 5 animals had hairloss outside the test site. Mineral oil, USP, and deionized water did not remove the sticky residue.
Challenge controls:
- Challenge control animals (5 males/5 females) were untreated during the induction phase (naive control); they were treated during the challenge phase (100 % of test article).
- An α-Hexylcinnamaldehyde (HCA) positive control group consisting of ten HCA test and ten HCA control guinea pigs was included in this study. The HCA positive control animals were treated with the HCA receiving 5 % w/v HCA in ethanol for induction and 2.5 % and 1.0 % w/v HCA in acetone for challenge. HCA control animals were treated only the challenge (also with 2.5 % and 1.0 % w/v HCA in acetone).
Positive control substance(s):
yes
Remarks:
α -Hexylcinnamaldehyde (HCA)

Results and discussion

Positive control results:
Following challenge with 2.5% w/v HCA in acetone, dermal scores of 1 were noted in 2/10 HCA test animals at the 24- and 48-hour scoring intervals. Dermal reactions in the remaining HCA test animals were limited to scores of 0 and +. Dermal reactions in the HCA challenge control animals were limited to scores of 0. Group mean dermal scores were noted to be higher in the HCA test animals as compared with the HCA challenge control animals. Following challenge with 1.0 % w/v HCA in acetone, a dermal score of 1 was noted in 1/10 HCA test animals at the 24-hour scoring interval. Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ±. Dermal reactions in the HCA challenge control animals were limited to scores of 0. Group mean dermal scores were noted to be higher in the HCA test animals as compared with the HCA challenge control animals.

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Dermal reactions were limited to scores of 0 to ±. Group mean score was 0.1.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: Dermal reactions were limited to scores of 0 to ±. Group mean score was 0.1. .
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
Dermal reactions in the test animals were limited to scores of 0 to ±. Group mean score was 0.1.
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: Dermal reactions in the test animals were limited to scores of 0 to ±. Group mean score was 0.1..
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No dermal reactions.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 100% . No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No dermal reactions..
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No dermal reactions.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No dermal reactions..
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
2.5% HCA in acetone
No. with + reactions:
2
Total no. in group:
10
Clinical observations:
Dermal scores of 1 were noted in 2 animals; Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ± .
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: positive control. Dose level: 2.5% HCA in acetone. No with. + reactions: 2.0. Total no. in groups: 10.0. Clinical observations: Dermal scores of 1 were noted in 2 animals; Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ± ..
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
2.5% HCA in acetone
No. with + reactions:
2
Total no. in group:
10
Clinical observations:
Dermal scores of 1 were noted in 2 animals; Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ± .
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: positive control. Dose level: 2.5% HCA in acetone. No with. + reactions: 2.0. Total no. in groups: 10.0. Clinical observations: Dermal scores of 1 were noted in 2 animals; Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ± ..
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
1.0% HCA in acetone
No. with + reactions:
1
Total no. in group:
10
Clinical observations:
A dermal score of 1 was noted in 1 animal; Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ±.
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: positive control. Dose level: 1.0% HCA in acetone. No with. + reactions: 1.0. Total no. in groups: 10.0. Clinical observations: A dermal score of 1 was noted in 1 animal; Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ±..
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
1.0% HCA in acetone
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Dermal reactions in all HCA test animals were limited to scores of 0 and ±.
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: positive control. Dose level: 1.0% HCA in acetone. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: Dermal reactions in all HCA test animals were limited to scores of 0 and ±..
Reading:
1st reading
Hours after challenge:
24
Group:
other: HCA challenge control
Dose level:
2.5% and 1.0% HCA in acetone
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No dermal reactions.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: other: HCA challenge control. Dose level: 2.5% and 1.0% HCA in acetone. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No dermal reactions..
Reading:
2nd reading
Hours after challenge:
48
Group:
other: HCA challenge control
Dose level:
2.5% and 1.0% HCA in acetone
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No dermal reactions.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: other: HCA challenge control. Dose level: 2.5% and 1.0% HCA in acetone. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No dermal reactions..

Any other information on results incl. tables

Test group and challenge control group

Following challenge with 100% (as received) OS131997H, IX.5 LT, dermal reactions in the test animals were limited to scores of 0 to ±. Dermal reactions in the challenge control animals were limited to scores of 0. Group mean dermal scores were noted to be slightly higher in the test animals as compared with the challenge control animals.

The sensitisation study animals gained weight during the test period and generally appeared in good health.

Table 3. Individual Challenge Data (Test material and Challenge Control)

Dermal scores

Group

Sex

24 hours*

48 hours*

Test material

(100 %)

M

0

0

M

0TSSY

0

M

0

0

M

0

0

M

±TSSY

±TSSY

M

0

0

M

0

0

M

0

0

M

0TSSY

0

M

0

0

F

0

0

F

0

0

F

0

0

F

0TSSY

0

F

0

0

F

0

0

F

0

0

F

±ED-1

±ED-1

F

0TSSY

0TSSY

F

0

0

MEAN

0.1

0.1

Challenge Control (100 %)

M

0

0

M

0

0

M

0

0

M

0TSSY

0

M

0

0

F

0

0

F

0

0

F

0

0

F

0TSSY

0

F

0

0

MEAN

0.0

0.0

Table 4: Individual Challenge Data (HCA Test and HCA Challenge Control)

Dermal scores

Group

Sex

2.5 % HCA

1.0 %

24 hours*

48 hours*

24 hours*

48 hours*

HCA Test

M

1DES

1

1ED-1

±

M

±

±

±

±

M

±

0

0

0

M

±

±

0

0

M

±

±

0

0

F

1

1

±

±

F

±

±

0

0

F

±

0

±

0

F

±

0

0

0

F

±IT

±

0

0

Mean

0.6

0.5

0.3

0.2

HCA Challnege Control

M

0

0

0

0

M

0

0

0

0

M

0

0

0

0

M

0

0

0

0

M

0

0

0

0

F

0IT

0

0

0

F

0

0

0

0

F

0IT

0

0

0

F

0DES

0DES

0DES

0DES

F

0IT

0

0

0

Mean

0.0

0.0

0.0

0.0

* Test site staining did not interfere with scoring;

TSSY = Test site staining yellow;

ED-1 Edema-Grade 1;

DES - Desquamation;

IT - Dermal irritation outside o f the test site.

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Test material is not considered to be a contact sensitiser in guinea pigs. The results of the HCA positive control study demonstrated that a valid test was performed and indicated that the test design would detect potential contact sensitisers.
Executive summary:

The dermal sensitisation potential of the test material (CAS No. 91648 -65 -6) was evaluated in Hartley-derived albino guinea pigs following OECD Guideline 406, EPA OPPTS 870.2600 and EU Method B.6 (Rodabaugh, 2006). Ten male and ten female guinea pigs were topically treated with 100 % (as received) test material once per week, for three consecutive weeks. Following a 2-week rest period, a challenge was performed whereby the 20 test and 10 previously untreated (naive) challenge control guinea pigs were topically treated with 100 % (as received) test material. Challenge responses in the test animals were compared with those of the challenge control animals. An α -Hexylcinnamaldehyde (HCA) positive control group consisting of ten HCA test and ten HCA control guinea pigs was included in this study. The animals were treated as above with the HCA test animals receiving 5 % w/v HCA in ethanol for induction and 2.5 % and 1.0 % w/v HCA in acetone for challenge.

Following challenge with 100 % test material, dermal reactions in the test animals were limited to scores of 0 to ±. Dermal reactions in the challenge control animals were limited to scores of 0. Group mean dermal scores were noted to be slightly higher in the test animals as compared with the challenge control animals. Following challenge with 2.5 % w/v HCA in acetone, dermal scores of 1 were noted in 2/10 HCA test animals at the 24- and 48-hour scoring intervals. Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ±. Dermal reactions in the HCA challenge control animals were limited to scores of 0. Group mean dermal scores were noted to be higher in the HCA test animals as compared with the HCA challenge control animals. Following challenge with 1.0 % w/v HCA in acetone, a dermal score of 1 was noted in 1/10 HCA test animals at the 24-hour scoring interval. Dermal reactions in the remaining HCA test animals were limited to scores of 0 and ±. Dermal reactions in the HCA challenge control animals were limited to scores of 0. Group mean dermal scores were noted to be higher in the HCA test animals as compared with the HCA challenge control animals.

Based on the results and evaluation criteria of this study, test material is not considered to be a contact sensitiser in guinea pigs. The results of the HCA positive control study demonstrated that a valid test was performed and indicated that the test design would detect potential contact sensitisers.