Registration Dossier

Administrative data

Description of key information

Weight of Evidence: Substance Skin Sensitisation: not sensitisng, 2017

Read-Across - methyl-2-[[(2,4-dimethyl-3-cyclohexen-1-yl)methylene]amino]benzoate: not sensitising up to 100% v/v and with induction in ethanol, Buehler test, OECD TG 406, 1985

Read-Across - methyl 2-[[[2,4(or 3,5)-dimethyl-3-cyclohexen-1-yl]methylene]amino]benzoate: not sensitising up to 100% v/v and with induction in ethanol, Guinea Pig OET test, 1985

Read-Across - oxydipropanol: not sensitising up to 100% v/v, Buehler test, US EPA OPP 81-6, 1995

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:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
1985
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP study following a method equivalent to a recognised guideline with acceptable deviations.
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included in attachment to IUCLID section 13.

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The read-across is based on the hypothesis that the source and target substances have similar physico-chemical, toxicological properties and because of common breakdown products and therefore potential mechanisms of action. The read-across is assessed as part of a weight of evidence approach. Further information is included in attachment to IUCLID section 13.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The source and target chemicals have comparable composition. Further information is included in attachment to IUCLID section 13

3. ANALOGUE APPROACH JUSTIFICATION
The source substance is a common degradant of the target substance and with common degradants to the target under physiological conditions. The read-across in weight of evidence indicates despite a common potential mode-of-action between source and target, no adverse effects are seen in several studies. When the information is taken together by expert judgement the weight of evidence indicates that there is an absence of adverse effects associated with the target.

4. DATA MATRIX
Further information is included in attachment to IUCLID section 13.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
yes
Remarks:
Only 10 animals used within the study, guideline recommends 20.
Principles of method if other than guideline:
Method employed in this study for the detection of delayed contact hypersensitivity was the guinea-pig test described by Buehler, E.V., Arch. Dermatol. 91, 171, (1965) and updated by Buehler, E.V. and Ritz, H.L., Current Concepts in Cutaneous Toxicity, pp. 215-40, (1980). Applicant accessment indicates that the scoring appears consistent with: Buehler, E.V. and Griffin F. Animal Models Dermatol., 55, (1975)
GLP compliance:
yes
Type of study:
Buehler test
Justification for non-LLNA method:
The test was conducted prior to 11 October 2016 as stated under Commission Regulation (EU) 2016/1688 and is used by read-across and as part of a Weight of Evidence adaptation under Regulation (EC) 1907/2006: Annex XI: section 1.5 read-across and Annex XI: section 1.2 weight of evidence.
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Recognised supplier
- Weight at study initiation: 417 - 460g (test group); 392-480g (control)
- Housing: In groups of five in grid bottomed polypropylene cages
- Diet (e.g. ad libitum): vitamin-C enriched pelleted diet ad libitum
- Acclimation period: 17 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17- 25°C
- Humidity (%): Not reported.
- Air changes (per hr): Air conditioned room (air changes not reported).
- Photoperiod (hrs dark / hrs light): 12 hours light / 12 hours dark
Route:
epicutaneous, occlusive
Vehicle:
other: ethanol
Concentration / amount:
Preliminary irritation testing: topical (occlusive): 12.5, 25, 50%v/v (ethanol) and 100% (undiluted)
Induction: - Topical: 100% (undiluted)
Challenge: - Topical: 100% (undiluted) and 50%v/v (ethanol)
Adequacy of induction:
highest technically applicable concentration used
Route:
epicutaneous, occlusive
Vehicle:
other: ethanol
Concentration / amount:
Preliminary irritation testing: topical (occlusive): 12.5, 25, 50%v/v (ethanol) and 100% (undiluted)
Induction: - Topical: 100% (undiluted)
Challenge: - Topical: 100% (undiluted) and 50%v/v (ethanol)
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
Test group: 10
Details on study design:
RANGE FINDING TESTS:
Preliminary Investigations: The intradermal and topical irritancy of a range of dilutions of the test substance was investigated.Four concentrations of the test material (100%, 50%, 25% and 12.5%) were used. The dilutions were prepared using ethanol. Each concentration was applied to one of four sites. The results of this dose ranging study indicated that the undiluted material was unlikely to act as a primary irritant at up to 100% (undiluted) concentration.

MAIN STUDY
A. INDUCTION EXPOSURE
Topical induction: A 0.5ml aliquot of neat test substance dorsal area was applied to the clipped left shoulder for 6 hours under occlusion (a 20mm square of surgical lint covered with surgical tape and elastic adhesive). Induction applications were made on days 1, 8 and 15. Controls were treated with ethanol vehicle only. This dosing procedure was repeated at weekly intervals on days eight and fifteen.

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: On day 28 of the study, 6-hour occluded challenge applications were made to the right flank with 50%v/v and 100% (undiluted) test item using (a 20mm square of surgical lint covered with surgical tape and elastic adhesive).
- Exposure period: 6 hours
- Test groups: 100% and 50%v/v
- Control group: 0% (vehicle only)
- Evaluation (hr after challenge): 24 and 48 hours.
Challenge controls:
Previously naive test group (treated only with vehicle) was treated with the test item as a challenge control at 50%v/v (in ethanol) and 100% (undiluted).
Positive control substance(s):
no
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No significant clinical signs, all gained bodyweight
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: 10.0. Clinical observations: No significant clinical signs, all gained bodyweight.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
50%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No significant clinical signs, all gained bodyweight
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 50%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No significant clinical signs, all gained bodyweight.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No significant clinical signs, all gained bodyweight
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No significant clinical signs, all gained bodyweight.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
50%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No significant clinical signs, all gained bodyweight
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 50%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No significant clinical signs, all gained bodyweight.

Table 1. Dermal reactions elicited in challenge application

Group

 

100%

 

50%

 

 

No. +ve reactions

 

 

24 hour

48 hour

24 hour

48 hour

 

 

Test

1*

0

0

0

0

 

0

 

2

0

0

0

0

 

0

 

3

0

0

0

0

 

0

 

4

0

0

0

0

 

0

 

5

0

0

0

0

 

0

 

6

0

0

0

0

 

0

 

7

0

0

0

0

 

0

 

8

0

0

0

0

 

0

 

9

0

0

0

0

 

0

 

10

0

0

0

0

 

0

Control

11

0

0

0

0

 

0

 

12

0

0

0

0

 

0

 

13

0

0

0

0

 

0

 

14

0

0

0

0

 

0

 

15

0

0

0

0

 

0

 

16

0

0

0

0

 

0

 

17

0

0

0

0

 

0

 

18

0

0

0

0

 

0

 

19

0

0

0

0

 

0

 

20

0

0

0

0

 

0

* Due to a technical error number 1 - occlusive dressing remained in position for 24 hours rather than 6 hours.

Interpretation of results:
GHS criteria not met
Conclusions:
Under the conditions of this study used, the test material is not considered to be a contact sensitizer.
Executive summary:

The study was performed according to a method equivalent to guideline OECD TG 406 consistent with Buehler method under GLP to assess the skin sensitisation potential of the test substance. The study was conducted using a preliminary irritation screen, an induction phase and a challenge phase. Preliminary irritation testing was for use in the induction phases of the study and the challenge phase of the study. Two groups, each of ten animals, were used for the study which was divided into an induction stage and a challenge stage. The induction stage consisted of three topical applications of the test material to the left shoulder of each of the ten test animals. These applications were made on Days one, eight and fifteen and each application lasted for six hours. Fourteen days after completion of the induction stage, all animals were challenged with two concentrations of the test material that had shown no evidence of irritation during the dose ranging study. No visible response was exhibited by any animal of the test or control group when challenged with the undiluted material and a 50% concentration of the material in ethanol. Under the conditions of this study, the test substance is not considered to be a contact sensitizer.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
1981
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
other: Non-GLP study following a methodology similar or equivalent to a scientifically recognised method with acceptable deviations
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included in attachment to IUCLID section 13.

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The read-across is based on the hypothesis that the source and target substances have similar physico-chemical, toxicological properties and because of common breakdown products and therefore potential mechanisms of action. The read-across is assessed as part of a weight of evidence approach. Further information is included in attachment to IUCLID section 13.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The source and target chemicals have comparable composition. Further information is included in attachment to IUCLID section 13

3. ANALOGUE APPROACH JUSTIFICATION
The source substance is a common degradant of the target substance and with common degradants to the target under physiological conditions. The read-across in weight of evidence indicates despite a common potential mode-of-action between source and target, no adverse effects are seen in several studies. When the information is taken together by expert judgement the weight of evidence indicates that there is an absence of adverse effects associated with the target.


4. DATA MATRIX
Further information is included in attachment to IUCLID section 13.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Principles of method if other than guideline:
Method was consistent or similar to the following literature method descriptions: Klecak, G. et al., J. Soc. Cosmet. Chem., vol. 28: 53-64 (1977);
Klecak, G. et al., Dermatotoxicology; 2nd ed., 200-219 (1982) and; Klecak, G. et al., Curr. Probl. Derm., vol. 14, 152-171 (1985)
GLP compliance:
no
Type of study:
open epicutaneous test
Justification for non-LLNA method:
The test was conducted prior to 11 October 2016 as stated under Commission Regulation (EU) 2016/1688 and is used by read-across and as part of a Weight of Evidence adaptation under Regulation (EC) 1907/2006: Annex XI: section 1.5 read-across and Annex XI: section 1.2 weight of evidence.
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Recognised supplier
- Females (if applicable) nulliparous and non-pregnant: Not specified.
- Weight at study initiation: 300 - 450g (test group and control group)
- Housing: In groups of three, solid floored polypropylene cages furnished with softwood sawdust
- Diet (e.g. ad libitum): vitamin-C enriched, recognised supplier pelleted diet, ad libitum
- Water (e.g. ad libitum): mains water supplier, ad libitum
- Acclimation period: Not reported.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ±3 °C
- Humidity (%): Not reported.
- Air changes (per hr): 20 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours light / 12 hours dark
Route:
epicutaneous, open
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% test item v/v / volume = 0.1 mL
Day(s)/duration:
21 days induction
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
epicutaneous, open
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% test item v/v in ethanol / volume = 0.025 mL
Day(s)/duration:
challenge: day 21 / 24 hours with observations at day 22, 23 and 24 and challenge: day 35 / 24 hours with observations at day 36, 37 and 38
Adequacy of challenge:
other: Minimal irritant concentration
Route:
epicutaneous, open
Vehicle:
other: ethanol
Concentration / amount:
30% test item v/v in ethanol / volume = 0.025 mL
Day(s)/duration:
challenge: day 21 / 24 hours with observations at day 22, 23 and 24 and challenge: day 35 / 24 hours with observations at day 36, 37 and 38
Adequacy of challenge:
highest non-irritant concentration
Route:
epicutaneous, open
Vehicle:
other: ethanol
Concentration / amount:
10% test item v/v in ethanol / volume = 0.025 mL
Day(s)/duration:
challenge: day 21 / 24 hours with observations at day 22, 23 and 24 and challenge: day 35 / 24 hours with observations at day 36, 37 and 38
Route:
epicutaneous, open
Vehicle:
other: ethanol
Concentration / amount:
3% test item v/v in ethanol / volume = 0.025 mL
Day(s)/duration:
challenge: day 21 / 24 hours with observations at day 22, 23 and 24 and challenge: day 35 / 24 hours with observations at day 36, 37 and 38
No. of animals per dose:
6 individuals per dose level and 6 within the control group
Details on study design:
RANGE FINDING TESTS:
Preliminary Investigations: The topical preliminary irritancy of a range of dilutions of the test substance was investigated. Five concentrations of the test material (100%, 30%, 10% 3% and 1%) were used. The dilutions were prepared using ethanol. Each concentration was applied to different sites. The results of this dose ranging study indicated that 100% (undiluted) concentration was the minimal irritant concentration (score = 1) and 30% was the maximum non-irritant concentration.

MAIN STUDY
A. INDUCTION EXPOSURE
Topical induction: A 0.1ml aliquot of neat test substance 100%v/v at 0.1 mL volume were applied dorsal area was applied daily for 21 consecutive days. Controls were treated with ethanol vehicle only 0.1 mL volume. This dosing procedure was repeated daily. In number 17, cracking appeared in the test group and therefore test item was applied at a different site.

B. CHALLENGE EXPOSURE
- No. of exposures: 2 (day 21 and re-challenge at day 36)
- Day(s) of challenge: On day 21 of the study, 24-hour open percutaneous challenge applications were made in the contralateral site.
- Exposure period: 24 hours
- Test groups: 100% (undiluted), 30%, 10% and 3%v/v in ethanol
- Control group: 100% (undiluted), 30%, 10% and 3%v/v in ethanol
- Evaluation (hr after challenge): 24, 48 and 72 hours
Challenge controls:
A naive control was similarly challenged to the test group which had not been previously exposed to the test item (induction with ethanol vehicle only).
Positive control substance(s):
no
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100% v/v
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
Minor dermal irritation observed (score = 1) in comparable range to vehicle control group challenge (score = 1)
Remarks on result:
no indication of skin sensitisation
Remarks:
observation periods: 24, 48 and 72 hours
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
30% v/v in ethanol
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
None.
Remarks on result:
no indication of skin sensitisation
Remarks:
observation periods: 24, 48 and 72 hours
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
10% v/v in ethanol
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Remarks:
observation periods: 24, 48 and 72 hours
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
3% v/v in ethanol
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Remarks:
observation periods: 24, 48 and 72 hours
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
100% v/v
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
Minor dermal irritation observed (score = 2 or 1) in comparable range to vehicle control group challenge (score = 2 or 1)
Remarks on result:
no indication of skin sensitisation
Remarks:
observation periods: 24, 48 and 72 hours
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
30% v/v in vehicle
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
Minor detmal irritation observed (score = 1) in comparable range to vehicle control group challenge (score = 1)
Remarks on result:
no indication of skin sensitisation
Remarks:
observation periods: 24, 48 and 72 hours
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
10% v/v in ethanol
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Remarks:
observation periods: 24, 48 and 72 hours
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
3% v/v in ethanol
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Remarks:
observation periods: 24, 48 and 72 hours

1. No reactions were observed at the day 21 challenge in any of the test or control animals at the maximal non-irritant concentration (i.e. 30%). However, scattered mild redness (score 1) was apparent at the minimal irritant concentration (i.e. 100%) in 2/6 test animals and 2/6 control animals. This was in agreement with the results obtained in the preliminary siting test.

2. In the day 35 challenge 1/6 animals in the test group and 2/6 animals in the control group produced reactions (Score = 1) at the maximal non-irritant concentration (i.e. 30%). In addition 6/6 animals in both the test and control groups showed reactions (Score = 2 or 1) at the minimal irritant concentration (100% v/v). Because reactions were comparable in both the test and control group animals at the day 35 challenge, these effects were associated with primary cutaneous irritation.

 

Applicant assessment indicates: The vehicle control group was not previously induced by the test item during the 21 day induction. All the test groups scores were within the irritation scoring seen in the vehicle control group treated with ethanol alone and then challenged at 100% and 30% v/v. No effects were reported in the open percutaneous test at 10% v/v or 3% v/v. All scores were within the range of scores seen during the test item induction phase of the test group with 100% test item application.

Interpretation of results:
GHS criteria not met
Conclusions:
Under the conditions of this study used, the test material is not considered to be a contact sensitizer.
Executive summary:

The study was performed according to a method equivalent to the Guinea Pig Open Epicutaneous Test detailed in Klecak et al (1977) to assess the skin sensitisation potential of the test substance. The topical preliminary irritancy of a range of dilutions of the test substance was investigated in a preliminary test. Five concentrations of the test material 100%, 30%, 10%, 3% and 1% were used. The dilutions were prepared using ethanol. Each concentration was applied to different sites. The results of this dose ranging study indicated that 100% (undiluted) concentration was the minimal irritant concentration (score = 1) and 30% was the maximum non-irritant concentration. Subsequently, induction was conducted using test material 100%, 30%, 10% and 3% v/v in ethanol. During induction 100%v/v test group indicated irritation during days 8 to 20 and cracking (maximum score 2). No irritation was seen in the vehicle group. The challenge phase commenced on day 21 using 100%, 30%, 10% and 3% v/v in ethanol. No reactions were observed at the day 21 challenge in any of the test or control animals at the maximal non-irritant concentration (i.e. 30%). However, scattered mild redness (score 1) was apparent at the minimal irritant concentration (i.e. 100%) in 2/6 test animals and 2/6 control animals. This was in agreement with the results obtained in the preliminary siting test. In the day 35 challenge 1/6 animals in the test group and 2/6 animals in the control group produced reactions (Score = 1) at the maximal non-irritant concentration (i.e. 30%). In addition 6/6 animals in both the test and control groups showed reactions (Score = 2 or 1) at the minimal irritant concentration (100% v/v). Because reactions were comparable in both the test and control group animals at the day 35 challenge, these effects were associated with primary cutaneous irritation. Under the conditions of the study it was considered that the test item was not sensitising by the open epicutaneous test.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
1995
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
other: GLP study following a method equivalent to a recognised guideline with acceptable deviations.
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included in attachment to IUCLID section 13.

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The read-across is based on the hypothesis that the source and target substances have similar physico-chemical, toxicological properties and because of common breakdown products and therefore potential mechanisms of action. Specifically, this source substance is a constituent of the source and target. With no indications of adverse effects. The read-across is assessed as part of a weight of evidence approach. Further information is included in attachment to IUCLID section 13.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The source and target chemicals have comparable composition. Further information is included in attachment to IUCLID section 13

3. ANALOGUE APPROACH JUSTIFICATION
The source substance is source substance is a constituent of the source and target. The read-across in weight of evidence indicates despite a common potential mode-of-action between source and target, no adverse effects are seen in several studies. When the information is taken together by expert judgement the weight of evidence indicates that there is an absence of adverse effects associated with the target.

4. DATA MATRIX
Further information is included in attachment to IUCLID section 13.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Qualifier:
according to
Guideline:
EPA OPP 81-6 (Skin Sensitisation)
Deviations:
no
Remarks:
Only 10 animals used within the study, guideline recommends 20. Only 5 naive organisms within negative control group utilised
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
yes
Remarks:
see above
GLP compliance:
yes
Type of study:
Buehler test
Justification for non-LLNA method:
The test was conducted prior to 11 October 2016 as stated under Commission Regulation (EU) 2016/1688 and is used by read-across and as part of a Weight of Evidence adaptation under Regulation (EC) 1907/2006: Annex XI: section 1.5 read-across and Annex XI: section 1.2 weight of evidence.
Species:
guinea pig
Strain:
not specified
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Recognised supplier
- Age at study initiation: Young adult
- Weight at study initiation: males: 386 to 455 g; females: 355 to 422 g
- Housing: Individually housed under standard laboratory conditions.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): municipal water ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature(ºC): 20 - 23 ºC
- Humidity (%): 35-65%
- Photoperiod (hrs dark / hrs light): 12 hours light / 12 hours dark
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% v/v / 0.5 mL
Day(s)/duration:
day 0, 6 and 14 in three 6 hour exposures
Adequacy of induction:
highest technically applicable concentration used
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% v/v / 0.1 mL
Day(s)/duration:
day 28 in 6 hour exposure
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
Test group: 10 (7m/3f)
Control group: 5 (3m/2f)
Details on study design:
RANGE FINDING TESTS:
To determine the correct concentrations to be used for induction (minimal irritation) and challenge (non-irritating), 5 guinea pigs were exposed to 4 concentrations (10, 25, 50 and 100% w/w in distilled water) of the test material to determine a non-irritating concentration and/or minimal irritation concentration. Locations of the applications alternated to variations; one test site had normal saline applied for comparative purposes. The substance was tested at 0, 10, 25, 50 and 100% w/w concentration in distilled water. There was no irritation and the 100% concentration of the test substance was seleceted for induction and challenge.

MAIN STUDY
A. INDUCTION EXPOSURE
- Test groups: 1
- Control group: 1
- Site: 1 per dose level
- Frequency of applications: Repeated, three times for 6 hours over a two week period (day 1, 8 and 14)
- Duration: 6 hours.
- Concentrations: 100%v/v (undiluted): A dose of 0.5 ml of the freshly prepared test solution was applied to a non-adherent sterile (aproximately 6 cm2) pad coverd by hypo-allerginec cloth tape. The pad was placed on the shaved surface of the animal, and then further occluded with a latex wrap to ensure adequate contract of the test substance with the skin. Then left untreated for approximately 2 weeks prior to primary challenge.

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: day 28.
- Exposure period: 6 hours
- Test groups: 1
- Control group: 1
- Site: 1 per dose level.
- Concentrations: Challanged at a virgin site (left side) with 0.5 ml at the non-irritating concentration at each site (at 100%v/v undiluted).
- Evaluation (hr after challenge): 24, 48 and 72 hours
Challenge controls:
A group of 5 naive animals were tested for comparison using the challenge concentration. A further naive group was tested for re-challenge where applicable.
Positive control substance(s):
no
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No systemic effects were observed and body weight gains were normal.
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: 10.0. Clinical observations: No systemic effects were observed and body weight gains were normal..
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No systemic effects were observed and body weight gains were normal.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No systemic effects were observed and body weight gains were normal..
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No systemic effects were observed and body weight gains were normal.
Remarks on result:
other: Reading: other: 3rd reading. . Hours after challenge: 72.0. Group: test group. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No systemic effects were observed and body weight gains were normal..
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
100% (previously naive; challenged with test item)
No. with + reactions:
1
Total no. in group:
5
Clinical observations:
Transient slight patchy erythema in one site
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 100% (previously naive; challenged with test item). No with. + reactions: 1.0. Total no. in groups: 5.0. Clinical observations: Transient slight patchy erythema in one site.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
100% (previously naive; challenged with test item)
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
None
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 100% (previously naive; challenged with test item). No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: None.
Reading:
other:
Hours after challenge:
72
Group:
negative control
Dose level:
100% (previously naive; challenged with test item)
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
None
Remarks on result:
other: Reading: other:. . Hours after challenge: 72.0. Group: negative control. Dose level: 100% (previously naive; challenged with test item). No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: None.
Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Under the conditions of this study used, the test material is not considered to be a contact sensitizer.
Executive summary:

The study was performed according to US EPA OPP 81-6 consistent with Buehler method under GLP to assess the skin sensitisation potential of the test substance. The study was conducted using a preliminary irritation screen, an induction phase and a challenge phase. Preliminary irritation testing was for use in the induction phases of the study and the challenge phase of the study. Two groups, one of ten animals for the test group was and 5 for the negative control were used for the study which was divided into an induction stage and a challenge stage. The induction stage consisted of three topical applications of the neat test material. These applications were made on day one, eight and fifteen and each application lasted for six hours. Fourteen days after completion of the induction stage, all animals were challenged with 100% (undiluted) test substance that had shown no evidence of irritation during the preliminary irritation test. No visible response was exhibited in the test group when challenged with the undiluted material. In the control group a response of score = 1 was observed that was transient and fully reversed within 48 hours which could be by applicant assessment to be a mild irritation response. Under the conditions of this study, the test substance is not considered to be a contact sensitizer.

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

Read-Across - methyl-2-[[(2,4-dimethyl-3-cyclohexen-1-yl)methylene]amino]benzoate: The study was performed according to a method equivalent to guideline OECD TG 406 consistent with Buehler method under GLP to assess the skin sensitisation potential of the test substance. The study was conducted using a preliminary irritation screen, an induction phase and a challenge phase. Preliminary irritation testing was for use in the induction phases of the study and the challenge phase of the study. Two groups, each of ten animals, were used for the study which was divided into an induction stage and a challenge stage. The induction stage consisted of three topical applications of the test material to the left shoulder of each of the ten test animals. These applications were made on Days one, eight and fifteen and each application lasted for six hours. Fourteen days after completion of the induction stage, all animals were challenged with two concentrations of the test material that had shown no evidence of irritation during the dose ranging study. No visible response was exhibited by any animal of the test or control group when challenged with the undiluted material and a 50% concentration of the material in ethanol. Under the conditions of this study, the test substance is not considered to be a contact sensitizer.

 

Read-Across - methyl 2-[[[2,4(or 3,5)-dimethyl-3-cyclohexen-1-yl]methylene]amino]benzoate: The study was performed according to a method equivalent to the Guinea Pig Open Epicutaneous Test detailed in Klecak et al (1977) to assess the skin sensitisation potential of the test substance. The topical preliminary irritancy of a range of dilutions of the test substance was investigated in a preliminary test. Five concentrations of the test material 100%, 30%, 10%, 3% and 1% were used. The dilutions were prepared using ethanol. Each concentration was applied to different sites. The results of this dose ranging study indicated that 100% (undiluted) concentration was the minimal irritant concentration (score = 1) and 30% was the maximum non-irritant concentration. Subsequently, induction was conducted using test material 100%, 30%, 10% and 3% v/v in ethanol. During induction 100%v/v test group indicated irritation during days 8 to 20 and cracking (maximum score 2). No irritation was seen in the vehicle group. The challenge phase commenced on day 21 using 100%, 30%, 10% and 3% v/v in ethanol. No reactions were observed at the day 21 challenge in any of the test or control animals at the maximal non-irritant concentration (i.e. 30%). However, scattered mild redness (score 1) was apparent at the minimal irritant concentration (i.e. 100%) in 2/6 test animals and 2/6 control animals. This was in agreement with the results obtained in the preliminary siting test. In the day 35 challenge 1/6 animals in the test group and 2/6 animals in the control group produced reactions (Score = 1) at the maximal non-irritant concentration (i.e. 30%). In addition 6/6 animals in both the test and control groups showed reactions (Score = 2 or 1) at the minimal irritant concentration (100% v/v). Because reactions were comparable in both the test and control group animals at the day 35 challenge, these effects were associated with primary cutaneous irritation. Under the conditions of the study it was considered that the test item was not sensitising by the open epicutaneous test.

 

Read-Across - oxydipropanol: The study was performed according to US EPA OPP 81-6 consistent with Buehler method under GLP to assess the skin sensitisation potential of the test substance. The study was conducted using a preliminary irritation screen, an induction phase and a challenge phase. Preliminary irritation testing was for use in the induction phases of the study and the challenge phase of the study. Two groups, one of ten animals for the test group was and 5 for the negative control were used for the study which was divided into an induction stage and a challenge stage. The induction stage consisted of three topical applications of the neat test material. These applications were made on day one, eight and fifteen and each application lasted for six hours. Fourteen days after completion of the induction stage, all animals were challenged with 100% (undiluted) test substance that had shown no evidence of irritation during the preliminary irritation test. No visible response was exhibited in the test group when challenged with the undiluted material. In the control group a response of score = 1 was observed that was transient and fully reversed within 48 hours which could be by applicant assessment to be a mild irritation response. Under the conditions of this study, the test substance is not considered to be a contact sensitizer.

 

The weight of evidence is based on read-across to the common degradant of the target substance, a structural analogue with a potential common mode-of-action and available data on a constituent of the target. None of the available studies indicates an adverse effect associated with skin sensitisation within the available studies when tested up to 100%v/v induction and challenged up to 100%v/v and in lower concentrations using ethanol vehicle. In the source substances the target test item can be considered to meet the skin irritation classification criteria for Regulation (EC) 1272/2008. There was no evidence in source substances of the test item causing enhanced responses (erythema or oedema) to the skin which might be indicative of skin sensitisation elicitation. All responses were within the vehicle control ranges. Additionally, the vehicle used potentially would have enhanced skin penetration of the common degradants. Additional effects were not observed. Based on the common degradants between the source and target there is no evidence of sensitisation with which to apply classification and labelling. There is indications that the test item will cause primary skin irritation meeting Regulation (EC) 1272/2008: skin irritation category 2: H315, based on the source and target data. Under the conditions of the available studies, the target substance weight of evidence is not skin sensitising.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The substance does not meet classification criteria under Regulation (EC) No 1272/2008 for skin sensitisation.

 

In the source substances the target test item can be considered to meet the skin irritation classification criteria for Regulation (EC) 1272/2008. There was no evidence in source substances of the test item causing enhanced responses (erythema or oedema) to the skin which might be indicative of skin sensitisation elicitation. All responses were within the vehicle control ranges. Additionally, the vehicle used potentially would have enhanced skin penetration of the common degradants. Additional effects were not observed. Based on the common degradants between the source and target there is no evidence of sensitisation with which to apply classification and labelling. There is indications that the test item will cause primary skin irritation meeting Regulation (EC) 1272/2008: skin irritation category 2: H315, based on the source and target data. Under the conditions of the available studies, the target substance weight of evidence is not skin sensitising.