bis(2-ethylhexyl) 4,4’-{6-[4-tert-butylcarbamoyl)anilino]-1,3,5-triazine-2,4-diyldiimino}dibenzoate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

June 1995

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1995

Report date:

1995

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

The guinea pig maximisation test is a standard test method that has been well validated. The study presented here was performed in 1995, well before guidelines preferring LLNA methods were introduced. As this existing study is sufficient to establish the skin sensitization potential of the test substance, further use of vertebrate animals to perform skin sensitization in unwarranted.

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

421-450-8

EC Name:

-

Cas Number:

154702-15-5

Molecular formula:

C44H59N7O5

IUPAC Name:

2-ethylhexyl 4-[(4-{[4-(tert-butylcarbamoyl)phenyl]amino}-6-[(4-{[(2-ethylhexyl)oxy]carbonyl}phenyl)amino]-1,3,5-triazin-2-yl)amino]benzoate

Details on test material:

- Name of test material (as cited in study report): RA 3643
- Physical state: white powder
- Lot/batch No.: 51/95
- Storage condition of test material: ambient and in the dark

In vivo test system

Test animals

Species:

guinea pig

Strain:

other: SPF-bred albino

Sex:

male/female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Wiga, Sulzfeld, Germany
- Age at study initiation: 2-3 weeks (ordered)
- Housing: in a mobile battery containing 4 cages; one cage per group of at most 10 animals
- Diet (e.g. ad libitum): pelleted, natural ingredient diet ad libitum
- Water (e.g. ad libitum): tap water ad libitum
- Acclimation period: 8 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+/- 3
- Humidity (%): 58-85%
- Air changes (per hr): about 10 air changes/hour
- Photoperiod (hrs dark / hrs light): 12 h light/12 h dark cycle

IN-LIFE DATES: From: May 31, 1995 To: June 8, 1995

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

petrolatum

Induction 2

Concentration / amount:

0.1 ml for the intradermal injection

Challenge

Challenge 1

Route:

intradermal and epicutaneous

Vehicle:

petrolatum

Challenge 2

Concentration / amount:

0.1 ml for the intradermal injection

No. of animals per dose:

Preliminary testr: two sets of 3 animals
Main study: test group of 5 males and 5 females; control group of 3males and 3 females

Positive control substance(s):

yes

Results and discussion

Positive control results:

A positive control study was performed to verify the lab could correctly identity dermal sensitizers. The positive control study used 5 animals as a negative control, and 10 animals (5 male and 5 female) as the test group. The positive control substance was DNCB.

The negative control animals were exposed to FCA and saline (1:1), maize oil, and maize oil and FCA (1:1) by injection at 6 sites per animal, and a topical application of vaseline in the induction phase.

For the test animals, they were exposed to FCA and saline (1:1), 0.1% test substance in maize oil, 0.1% test substance in maize oil and FCA (1:1) by injection, and a topical application of 0.1% test substance in vaseline.

For the challenge phase, all animals were exposed to a dermal application of 0.05% test substance in vaseline.

In the control group, only one animal showed signs of irritation after the challenge phase (score of 1), while all animals in the test group showed signs of irritation (scores of 1-3).

The results show the lab can successfully identify skin sensitizing substances.

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

0.05% DNCB in vaseline

No. with + reactions:

9

Total no. in group:

10

Clinical observations:

One animal was killed due to poor condition. All other animals in the positive control group showed skin irritation with scores of 0-3.

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

30% RA 3646 in vaseline

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

No signs of skin irritation were seen.

Results 3

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

Vaseline only

No. with + reactions:

0

Total no. in group:

6

Clinical observations:

No signs of skin irritation were seen.

Results 4

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

0.05% DNCB

No. with + reactions:

9

Total no. in group:

10

Clinical observations:

One animal in the positive control group showed poor condition and was killed. All other animals showed skin reaction of 1-3 during the challenge phase. Two animals had small scratches and another two animals showed scaliness.

Remarks on result:

positive indication of skin sensitisation

Results 5

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

0

No. with + reactions:

0

Total no. in group:

6

Clinical observations:

None

Remarks on result:

no indication of skin sensitisation

Results 6

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

30% dilution of test item

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

no dermal reaction

Remarks on result:

no indication of skin sensitisation

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

the test substance RA 3643 is considered not to be a sensitizer

Executive summary:

The test substance RA 3643 was examined for possible sensitizing properties by a maximization test in guinea pigs using 10 test animals and 6 controls.

The test comprised:

TEST ANIMALS

- induction treatment by intradermal injections of Freund's Complete Adjuvant (FCA) 1:1 diluted with isotonic saline, a 3% dilution of RA 3643 in maize oil, and a 3% dilution of RA 3643 in vaseline (test site pretreated with sodium lauryl sulfate),

- challenge treatment, 14 days after the last induction, by topical application of a 30% dilution of RA 3643 in vaseline and of vaseline alone,

CONTROLS

- induction treatment by intradermal injections of FCA 1:1 diluted with isotonic saline, maize oil, and FCA/saline 1:1 diluited with maize oil, followed one week later by topical application of patches loaded with vaseline (test sites pretreated with sodium lauryl sulfate), and

- challenge treatment, 14 days after the last induction, by topical application of a 30% dilution of RA 3643 in vaseline and of vaseline alone,

None of the test animals reacted positively. Therefore, the test substance RA 3643 is considered not to be a sensitizer. Since clear and unequivocal results were obtained in this study, no need for further examination in an additional batch of test and control animals is required.