SILATRIZOLE

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 13 November 1995 to 01 February 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study conducted in compliance with OECD Guideline 406 without any deviations.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

yes

Remarks:

Mixture of FCA:physiological saline was chosen instead of FCA only in order to decrease the site effects of FCA when applied alone

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

(16-06-1994)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Adequate data from non-LLNA method is already available

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

422-940-4

EC Name:

-

Cas Number:

155633-54-8

Molecular formula:

C24H39N3O3Si3

IUPAC Name:

2-(2H-1,2,3-benzotriazol-2-yl)-4-methyl-6-[2-methyl-3-(2,2,4,6,6-pentamethyl-3,5-dioxa-2,4,6-trisilaheptan-4-yl)propyl]phenol

Test material form:

other: solid

Details on test material:

- Name of test material (as cited in study report): G4375
- Physical state: whitish solid
- Analytical purity: 98% minimum
- Impurities (identity and concentrations): Methanol (<100 ppm) and Isopropanol (<100 ppm)
- Purity test date: certificate of analysis 20/10/1995
- Lot/batch No.: DEF/C 95003/B
- Date of reanalysis: september 1996
- Stability under test conditions: Stable under storage condition; stability unknown in corn oil / acetone 20/80 and in a 1:1 (v/v) mixture of FCA/physiological saline, therefore is excluded from the statement of compliance.
- Storage condition of test material: In the original container at room temperature away from direct sunlight.

In vivo test system

Test animals

Species:

guinea pig

Strain:

other: Himalayan white spotted (Ibm : GOHI ; SPF-quality guinea pigs)

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: BRL, Biological research Laboratories Ltd., Wölfertrasse 4, 4414 Füllinsdorf, Switzerland
- Age at study initiation: 5-7 weeks
- Weight at study initiation: Control and test group: 334-443 g; Range-finding test: 414-476 g
- Housing: Individually in Makrolon type-3 cages with autoclaved standard softwood bedding (“Lignocel”, Scill AG, CH-4132 Muttenz).
- Diet: Pelleted standard Nafag Ecosan 845 25W4, Batch n 94/95 and 108/95 guinea pig breeding/maintenance diet (“Nafag”, Nähr- und Futtermittel AG, CH-9202 Gossau), ad libitum.
- Water: community tap water from Itingen, ad libitum. Once weekly, additional supply of ascorbic acid (1g/l) via the drinking water.
- Acclimation period: One week for the control and test group. No acclimatization for the animals of the Range-finding study.


ENVIRONMENTAL CONDITIONS
- Temperature: 21 ± 3 ºC
- Humidity: 40-70%
- Air changes: 10-15 air changes per hour
- Photoperiod: 12 h dark / 12 h light

IN-LIFE DATES: From: 13 November 1995 To: 14 December 1995

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

other: Corn oil / acetone (20/80)

Induction 2

Concentration / amount:

Range finding test:
- Intradermal induction exposure: 1, 3 and 5%
- Topical induction exposure: 60, 70, 80 and 85%

Main test:
- Intradermal induction exposure: 5%
- Topical induction exposure: 85%
- Topical challenge exposure: 85%

Challenge

Challenge 1

Route:

epicutaneous, semiocclusive

Vehicle:

other: Corn oil / acetone (20/80)

Challenge 2

Concentration / amount:

Range finding test:
- Intradermal induction exposure: 1, 3 and 5%
- Topical induction exposure: 60, 70, 80 and 85%

Main test:
- Intradermal induction exposure: 5%
- Topical induction exposure: 85%
- Topical challenge exposure: 85%

No. of animals per dose:

- Range finding test: 6 males
- Main test: 10 males for negative control and 20 animals for treatment group

Details on study design:

RANGE FINDING TEST:
- Intradermal induction exposure: intradermal injections (0.1 ml/site) were made into the clipped flank of 2 animals at concentrations of 5, 3 and 1% of the test article in corn oil/acetone (20/80). The resulting dermal reactions were assessed 24 h later. A 5% dilution was selected for intradermal induction.
- Topical induction exposure: Both flanks of each of other 4 guinea pigs were clipped and shaved just prior to the application. The test substance at 60, 70, 80 and 85% in corn oil/acetone (20/80) was applied topically under 4 occlusive patches (covered by a strip of aluminum foil and firmly secured by elastic plaster and covered with impervious adhesive tape) per animal, saturated with the test article dilutions, and held in contact with the skin for 24 hours. The reaction sites were assessed 24 and 48 h after removal of the patches for erythema and oedema according to Draize scale. The 85% dilution was selected for the topical induction and challenge.

MAIN STUDY
A. INDUCTION EXPOSURE: INTRADERMAL
- No. of exposures: One
- Day of induction: Day 1
- Test group: 3 pairs of intradermal injections (0.1 mL each) of a mixture of 1:1 (v/v) of Freund’s Complete adjuvant (FCA) and physiological saline, test article at 5% in corn oil/acetone (20/80), test article at 5% in a 1:1 (v/v) of FCA and physiological saline.
- Control group: 3 pairs of intradermal injections (0.1 mL each) of a mixture of 1:1 (v/v) of FCA and physiological saline, corn oil/acetone (20/80), 1:1 (w/w) mixture of corn oil/acetone (20/80) and 1:1 (v/v) of FCA and physiological saline.
- Site: an area of dorsal skin from the scapular region (approximately 6x8cm) was clipped free of hair.

B. INDUCTION EXPOSURE: TOPICAL
- No. of exposures: One
- Day of induction: Day 8
- Exposure period: 48 h
- Test group: After a 24h-topical exposure to Sodium Lauryl sulfate (10% in paraffinum perliquidum), a occlusive patch (covered by a strip of aluminum foil and firmly secured by elastic plaster and covered with impervious adhesive tape) saturated with the test article (85% in corn oil/acetone (20/80)) was applied topically.
- Control group: After a 24h-topical exposure to Sodium Lauryl sulfate (10% in paraffinum perliquidum), a occlusive patch saturated with corn oil/acetone (20/80)) was applied topically.
- Site: Same intradermally injected area (an area of scapular region clipped free of hair)
- Frequency of applications: Single application
- Evaluation (h after removal of induction patch): 24 and 48 h


C. CHALLENGE EXPOSURE: TOPICAL
- No. of exposures: One
- Day of challenge: Day 22
- Exposure period: 24 h
- Test and control groups: Area of the left and right flanks of treated and control animals were clipped and cleared of hair and 2 occlusive patches (covered by a strip of aluminum foil and firmly secured by elastic plaster and covered with impervious adhesive tape) saturated with the test article (85% in corn oil/acetone (20/80)) (left flank) and the vehicle only (right flank) were applied topically.
- Site: left and right flanks treated with test item dilution and vehicle only, respectively.
- Evaluation (h after removal of challenge patch): 24 and 48 h

OTHER:
- Mortality/clinical signs (local and systemic): daily during observation period
- Body weight: at the beginning of the acclimatization period, at day one and at the termination of the test.
- A 24h-topical exposure to Sodium Lauryl sulfate (10% in paraffinum perliquidum) was performed before topical induction phase in order to induce skin irritation.
- The reaction sites were scored 24 and 48h after the removal of the patch for erythema and oedema according to Draize scale. The readings were made under artificial fluorescent light.

Challenge controls:

In challenge period, the control group was treated once with the vehicle and with the test item at 85% in vehicle.

Positive control substance(s):

yes

Remarks:

2-Mercaptobenzothiazol and α-hexylcinnamaldehyde

Results and discussion

Positive control results:

In recent reliability check, 15% 2-Mercaptobenzothiazol in mineral oil and 3% α-hexylcinnamaldehyde in PEG 400 (positive historical controls) were found to induce an appropriate response in guinea-pig (95% of the animals at 48 h and 72 h and 75% of the animals at 48h and 45% at 72h, respectively).

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

other: positive control

Hours after challenge:

48

Group:

positive control

Dose level:

15% 2-Mercaptobenzothiazol in mineral oil

No. with + reactions:

9

Total no. in group:

10

Remarks on result:

positive indication of skin sensitisation

Results 2

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

85% in corn oil/acetone (20/80)

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: 85% in corn oil/acetone (20/80). No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: none.

Results 3

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

85% in corn oil/acetone (20/80)

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: 85% in corn oil/acetone (20/80). No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: none.

Results 4

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

0 %

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: negative control. Dose level: 0 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Results 5

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

0 %

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: negative control. Dose level: 0 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Any other information on results incl. tables

- During the experimental period, none of the animals (control and treated groups) presented edema or erythema.

- No erythema or edema was observed after 48 and 72 h patch removal.

- One animal of the test group lost weight during the acclimatization period thus considered incidental.

- No symptoms of systemic toxicity were observed in the animals.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Remarks:

Criteria used for interpretation of results: EU

Conclusions:

Under these test conditions, Silatrizole (encoded "G4375") is not classified as skin sensitiser according to the Regulation (EC) N° 1272-2008 (CLP).

Executive summary:

In a GLP Magnusson & Kligman maximisation study (GPMT) performed according to OECD Guideline 406, 20 male Himalayan white spotted guinea pigs were induced with three pairs of intradermal injections (0.1 mL each) of a mixture of 1:1 (v/v) of Freund’s Complete adjuvant (FCA) and physiological saline, test article at 5% in corn oil/acetone (20/80), test article Silatrizole (encoded "G4375") at 5% in a 1:1 (v/v) of FCA and physiological saline, on Day 0 on three different sites on an area of dorsal skin from the scapular region.

Control group of 10 males was intradermally induced (0.1 mL each)with 1:1 (v/v) of FCA and physiological saline, corn oil/acetone (20/80), 1:1 (w/w) mixture of corn oil/acetone (20/80) and 1:1 (v/v) of FCA and physiological saline.

On Day 7, a topic application of 10% sodium lauryl sulfate solution in paraffimun perliquidum was performed on the same area of injections.

On Day 8, a topic application was performed using a occlusive patch saturated with the test article (85% in corn oil/acetone (20/80)) (experimental group) or corn oil/acetone (20/80) (control group) held in contact to the tested area for 48 hours.

After a rest period of two weeks, on Day 22, challenge exposure began: two occlusive patches saturated with the test article (85% in corn oil/acetone (20/80)) or corn oil/acetone (20/80) were applied on the left and right flank of the animals (test and control), respectively. The test concentration for the main study was determined from a range-finding study using 6 animals (2 for intradermal induction and 4 for topical application).

No impact was observed in the study, since no skin reactions were observed in the animals during the challenge period. No skin reactions were noted at the challenge sites of the test or control group animals at the 48 or 72 h observations. Silatrizole produced a 0 % (0/10) sensitization rate and was considered to be a non-sensitizer to guinea pig skin. Positive control (15% 2-Mercaptobenzothiazol in mineral oil and 3% α-hexylcinnamaldehyde in PEG 400) exhibited evidence of sensitization. 

 

Under these test conditions, Silatrizole is not classified as skin sensitizer according to the Annex VI to the Regulation (EC) N° 1272-2008 (CLP).