Reaction products of N,N'-ethane-1,2-diylbis(1,3-propanediamine), cyclohexane, peroxidized 4-butylamino-2,2,6,6-tetramethylpiperidine and 2,4,6-trichloro-1,3,5-triazine

Endpoint summary

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1997-02-03 - 1997-04-15

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline Study in compliance with GLP

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Qualifier:

according to guideline

Guideline:

EPA OPP 81-6 (Skin Sensitisation)

Qualifier:

according to guideline

Guideline:

EPA OTS 798.4100 (Skin Sensitisation)

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

A valid Guinea Pig Maximization Test (GPMT) is available with the test item and therefore no LLNA is required.

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male/female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Sprague Dawley, Inc, Indianapolis, IN 46229
- Age at study initiation: Young adult
- Weight at study initiation: 302 - 481 grams at the start of the study
- Housing: housing and animal care conformed to standards (National Research Council, 1996)
- Teklad Guinea Pig Diet: ad libitum
- Tap water: ad libitum
- Acclimation period: at least five days
- Experimental Start Date: February 4, 1997
- Experimental Termination Date: March 11, 1997

ENVIRONMENTAL CONDITIONS
- Temperature: 10 - 26 °C
- Humidity: 30 - 70 %
- Photoperiod (hrs dark / hrs light): 12/12

Route:

intradermal and epicutaneous

Vehicle:

other: diethyl phthalate

Concentration / amount:

Intradermal induction exposure: 0.5% (w/v) concentration of test item in diethyl phthalate
Epicutaneous induction exposure: 25% (w/v) concentration of test item in diethyl phthalate
Epicutaneous challenge exposure: 5% (w/v) concentration of test item in diethyl phthalate
(Test group exposures were based on pretest results)

Route:

epicutaneous, occlusive

Vehicle:

other: diethyl phthalate

Concentration / amount:

Intradermal induction exposure: 0.5% (w/v) concentration of test item in diethyl phthalate
Epicutaneous induction exposure: 25% (w/v) concentration of test item in diethyl phthalate
Epicutaneous challenge exposure: 5% (w/v) concentration of test item in diethyl phthalate
(Test group exposures were based on pretest results)

No. of animals per dose:

In total 50 animals were used:
Pretest:
- 8 topical pilot animals (4 males, 4 females)
- 2 intradermal injection pilot animals (1 male, 1 female)
Main test:
- 20 test animals (10 males, 10 females) [test group]
- 10 vehicle control animals (5 males, 5 females) [vehicle control group]
- 10 naive control animals (5 males, 5 females) [naive control group]

Details on study design:

RANGE FINDING TESTS:
- The irritation phase had the purpose of evaluating the irritation potential of test material at the levels to be used in the injection induction, topical induction, and challenge phase. The primary irritancy of the test material, when both injected and topically applied, was evaluated.

The procedure employed for these investigations was as follows:
Intradermal injections:
Intradermal injections (0.1 mL/site) were made into the clipped flank of two guinea-pigs at concentrations of 10%, 5%, 2.5%, 1%, 0.5%, 0.25%, 0.1%, and 0.05% (w/v) of the test item in diethyl phthalate. The resulting dermal reactions were assessed examined three days following injection.
- Epidermal applications:
Patches were endowed with a 0.4 mL quantity of 25%, 10%, 5%, 2.5%, 1%, 0.5%, 0.25%, and 0.1% (w/v) test item solution in diethyl phthalate and applied to the clipped and shaved flanks of each of eight guinea-pigs. Each animal was treated with four different concentrations; the position of the different concentrations on the animals was varied to adjust for possible site-to-site variation in response. Patches were secured by an elastic adhesive bandage wound around each of the animal's torso until overlapping.
The dressings were removed after an exposure period of 24 hours and the reaction sites were assessed for erythema and edema on a numerical basis.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures:
2 exposures, i.e. a single intradermal exposure in week 1 (6 injections/animal) and an epicutaneous exposure in week 2 (closed patch)
- Exposure period: single application (intradermal exposure), 48 hours (epicutaneous exposure)
- Control groups: 2 groups of 10 animals
- Frequency of applications: 1 intradermal application (week 1) and 1 epicutaneous application (week 2)
- Concentrations: 0.5 % (intradermal exposure, day 0), 25 % (epicutaneous exposure, day 7)
- Site: nape and upper back area

A1. INTRADERMAL INJECTIONS
- No. of exposures: 6 intradermal injections in groups of two per animal
- Test group (1 group of 20 animals):
1) 50% (v/v) FCA/distilled water emulsion
2) Test item as a 0.5% (w/v) formulation in diethyl phthalate
3) Test item as a 0.5% (w/v) formulation in FCA/distilled water emulsion (50% v/v)
- Control group (1 group of 10 animals - Vehicle Control):
1) 50% (v/v) FCA/distilled water emulsion
2) Undiluted diethyl phthalate
3) Diethyl phthalate as a 50% (w/v) formulation in FCA/distilled water emulsion (50% v/v)
- Site: nape and upper back area

A2. EPIDERMAL APPLICATIONS
- On Day 6, the nape and upper back area of each animal in the test and the vehicle control groups were re-clipped. On Day 7, a 0.8 mL quantity of the freshly prepared material was applied onto the previously described patch system. The test substance was applied to the test group as a 25% w/v formulation in diethyl phthalate. The undiluted vehicle material was applied to the vehicle control group. Patches were fixed as described above. The dressings were left in place for approximately 48 hours. Reaction sites were assessed for erythema and edema 24 and 48 hours after removal of the dressing.

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day of challenge: Day 21
- Exposure period: 24 hours
- Test group: On Day 20, the appropriate skin sites of the test, the naive control, and the vehicle control groups were clipped. On Day 21, a 0.4 mL quantity of the freshly prepared formulation was applied onto the described patch system. The test substance patch was applied to the clipped area of the left flank, and a vehicle material patch was applied to the clipped surface of the right flank of each animal in the test, naive control, and vehicle groups.
- Control groups (Vehicle Control Group, Naive Control Group): The control animals were treated in the same way as described above.
- Concentration: 5% (w/v) formulation in diethyl phthalate
- Evaluation: 24, 48, and 72 hours after removal of the dressing

OTHER:
Body Weights: Initial body weights were measured just prior to the first exposure of a respective group of animals, with the exception of naive control animals which were weighed concurrently with the induced groups of animals which they served. Final body weights were taken subsequent to the
receipt of the Sponsor's authorization to terminate testing. Final body weights were not taken for pilot animals due to their short term study participation.

Challenge controls:

No positive reactions were noted after the challenge application, neither when treated with diethyl phthalate alone nor when treated with a 5% test item formulation.

Positive control substance(s):

yes

Remarks:

Historical positive control data on Alpha-Hexylcinnamaldehyde, tech., 85% as a 5% (w/v) formulation in Acetone included in the report

Positive control results:

Historical positive control data available.

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

5%

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

none

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: none.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

5%

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

none

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: none.

Reading:

other: 3rd reading

Hours after challenge:

72

Group:

test chemical

Dose level:

5%

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

none

Remarks on result:

other: Reading: other: 3rd reading. . Hours after challenge: 72.0. Group: test group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: none.

Reading:

1st reading

Hours after challenge:

24

Group:

other: vehicle control group

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: other: vehicle control group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Reading:

2nd reading

Hours after challenge:

24

Group:

other: vehicle control group

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 24.0. Group: other: vehicle control group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Reading:

other: 3rd reading

Hours after challenge:

72

Group:

other: vehicle control group

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

other: Reading: other: 3rd reading. . Hours after challenge: 72.0. Group: other: vehicle control group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Reading:

1st reading

Hours after challenge:

24

Group:

other: naive control group

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: other: naive control group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Reading:

2nd reading

Hours after challenge:

48

Group:

other: naive control group

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: other: naive control group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Reading:

other: 3rd reading

Hours after challenge:

72

Group:

other: naive control group

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

other: Reading: other: 3rd reading. . Hours after challenge: 72.0. Group: other: naive control group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Interpretation of results:

not sensitising

Remarks:

Migrated information

Conclusions:

Under the test conditions chosen, the test article did not cause allergic reactions to guinea pig skin and is therefore not considered as skin sensitizer.

Executive summary:

In a guinea pig maximization test according to OECD guideline No. 406, 10 male and 10 female animals were first induced and then challenged with the test article to investigate its sensitization potential. Induction was a two-stage operation: First, three pairs of intradermal injections (0.5 % test substance in diethyl phthalate) were made into the nape and upper back area of the animals with a 1:1 mixture (v/v) of FCA/ physiological saline, the test substance in diethyl phthalate, and the test substance in a 1:1 mixture (v/v) of FCA/ physiological saline. One week later, 0.8 ml of the test article in diethyl phthalate at a concentration of 25% was spread onto a patch consisting of a 20 mm x 20 mm Webril® pad and applied to the clipped backs of the animals for 48 hours. Two weeks after the epidermal induction application the animals were tested on the flank with 5% test substance in diethyl phthalate and the vehicle alone for 24 hours. Twenty four hours after removing the dressings the challenge reactions were graded according the Draize scoring scale. Two additional evaluations were made 48 and 72 hours after removing the dressings. A control group (5 m/5 f) was treated with adjuvant and the vehicle during the induction period. A second control group remained untreated during induction. During the challenge period both control groups were treated with the vehicle as well as with the test compound. No animal of the test group was sensitized by the test substance, all skin reactions at any time point were scored 0. Therefore, under the experimental conditions of this study, the test material is non-sensitizing when topically applied to albino guinea pigs.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

The test substance was investigated for its sensitizing potential in a GLP-compliant Maximization test according to OECD guideline 406 (HILL TOP RESEARCH, Ltd., 97-8374-21, 1997). Dunkin-Hartley guinea pigs received intradermal induction treatments with 0.5% in diethyl phthalate and one epicutaneous induction treatment with 25% in diethyl phthalate. For intradermal induction, three pairs of injections (adjuvant/saline; test item in vehicle; test item in vehicle plus adjuvant/saline) were given. Epidermal challenge was performed by occlusive application for 24 h two weeks later (5% in diethyl phthalate). Concurrent negative vehicle control animals were treated similarly, except that during the induction phase the test substance was omitted. Furthermore, a naive control group was part of the challenge phase. No skin reactions were observed either for control or test group animals and therefore no animal was sensitized by the test article under the experimental conditions employed. The sensitivity of the strain is controlled every six month with alpha-Hexylcinnamaldehyde. In conclusion, the test substance is not considered to be a skin sensitizer in albino guinea pigs and does not require classification.

Migrated from Short description of key information:
In a dermal sensitization study male and female guinea pigs were tested with the test substance in a Maximization Test according to OECD TG 406. No sensitized animals were observed after treatment with the test substance in a concentration of 5 %. Under the conditions of this study, the test substance was not a skin sensitizer.

Justification for selection of skin sensitisation endpoint:
Guideline Study in compliance with GLP

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information:

no data

Migrated from Short description of key information:
no data

Justification for classification or non-classification

Classification, Labeling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for the purpose of classification under Regulation (EC) No.1272/2008. Based on the present data, classification for sensitization is not warranted under Regulation (EC) No.1272/2008.