Propanoic acid, 3-[[bis(2-methylpropoxy)phosphinothioyl]thio]-2-methyl-

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

9-NOV-1999 - 20-DEC-1999

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

Report date:

2000

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

(1993)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

The study was conducted in 2000, when the GPMT was an international accepted and recommended method to assess sensitizing potential.

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

434-070-2

EC Name:

-

Cas Number:

268567-32-4

Molecular formula:

C12 H25 O4 P S2

IUPAC Name:

3-{[bis(2-methylpropoxy)(sulfanylidene)-λ⁵-phosphanyl]sulfanyl}-2-methylpropanoic acid

Test material form:

liquid: viscous

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability under test conditions: Stable in PEG 400 and in a 1:1 (v/v) mixture of FCA/physiological saline for at least 2 h

In vivo test system

Test animals

Species:

guinea pig

Strain:

other: Ibm: GOHI; SPF-quality

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: RCC Ltd. Biotechnology & Animal Breeding Division, CH-4414 Füllinsdorf / Switzerland.
- Age at study initiation: 5 - 7 weeks.
- Weight at study initiation: 337 - 371 g.
- Housing: individually in Makrolon type-4cages.
- Diet: pelleted standard Nafag Ecosan 845 25W4, guinea pig breeding/maintenance diet (containing Vitamin C), ad libitum.
- Water: tap water, ad libitum.
- Acclimation period: 1 week.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 40 - 70
- Air changes (per hr): 10 - 15
- Photoperiod (hrs dark / hrs light): 12 / 12

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

polyethylene glycol

Concentration / amount:

50 %

Day(s)/duration:

day 0

Induction 2

Route:

epicutaneous, occlusive

Vehicle:

unchanged (no vehicle)

Concentration / amount:

0.3 mL; 100 %

Day(s)/duration:

day 8 / 48 h

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

polyethylene glycol

Concentration / amount:

50 %

Day(s)/duration:

day 21 / 24 h

No. of animals per dose:

Pretest: 3 animals.
Main test: 10 animals in treatment group, 5 animals in control group.

Details on study design:

RANGE FINDING TESTS:
Pretest:
Intradermal Injection:
Four intradermal injections (0.1 mL/site) of a 1:1 mixture of FCA/physiological saline were made in the shaved neck of one guinea pig. One week later intradermal injection (0.1 mL/site) was made into the clipped flank of the same guinea pig at concentrations of 100 %, 75 % and 50 % of the test article in PEG 400. Dermal reactions were assessed 24 h later. Based on the results a concentration of 50 % was selected for intradermal induction in the main study.
Epidermal Application:
Four intradermal injections (0.1 ml/site) of a 1:1 mixture of FCA/physiological saline were made in the shaved neck of two guinea pigs. One week later, 4 patches were saturated with the test article at concentrations of 100 %, 75 %, 50 % and 25 % in PEG 400, respectively. The patches were applied to the clipped and shaved flanks of each of the two animals under occlusive conditions. The exposure period was 24 h. After removal of the patches the skin was examined for signs of irritation.
According to results of the pretest for intradermal injection (induction) in the main study a concentration of 50 % of test article in vehicle was selected. Undiluted (100 %) test article was selected for epidermal induction, and 50 % in vehicle for epidermal challenge.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2;
Intradermal: three pairs of intradermal injections (1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline, the test article, at 50 % in PEG 400 and the test article at 50 % in a 1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline; 0.1 mL/site)
Epidermal: undiluted test article one week after intradermal injections.
- Exposure period: epidermal application was performed for 48 h.
- Test groups: 10 males.
- Control group: 5 males.
- Site: dorsal skin from the scapular region.

B. CHALLENGE EXPOSURE (TWO WEEKS AFTER INDUCTION)
- No. of exposures: 1.
- Day(s) of challenge: 1.
- Exposure period: 24 h.
- Test groups: 10 males.
- Control group: 5 males.
- Site: flank.
- Concentrations: 50 % test article in vehicle.
- Evaluation (hr after challenge): approximately 25 and 48 hours after challenge.

Challenge controls:

PEG 400

Positive control substance(s):

yes

Remarks:

2-Mercaptobenzothiazole (independently performed, report attached)

Results and discussion

Positive control results:

Validation of sensitivity of the Maximization-Test was performed from 1998-12-01 to 1999-01-08 (RCC Project 903407). In accordance to OECD 406, 2-Mercaptobenzothiazole was selected as a positive control. The intradermal induction of sensitization was performed with a 5 % dilution of the test article in mineral oil and in an emulsion of FCA/physiological saline. The epidermal induction of sensitization was conducted under occlusion with the test article at 50 % in mineral oil. Two weeks after the epidermal induction application the challenge was completed by epidermal application of the test article at 10 % in mineral oil under occlusive dressing. The animals of the control group were induced with mineral oil and FCA/physiological saline and challenged similarly to those of the test group. Cutaneous reactions, i.e. erythema and eschar, as well as edema formation were evaluated at 24 and 48 hours after removal of the dressing.
90 % (at the 24-hour reading) and 100 % (at the 48-hour reading) of the animals of the test group were observed with very slight to moderate/severe erythematous reactions after treatment with a non-irritant test article concentration of 10 % in mineral oil. No skin reactions were observed in the control group.

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

0 %

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

no skin irritations were observed

Results 2

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

50 %

No. with + reactions:

7

Total no. in group:

10

Clinical observations:

positive reactions consisted of discrete/patchy to intense erythema and swelling of the skin

Results 3

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

0 %

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

no skin irritations were observed

Results 4

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

50 %

No. with + reactions:

7

Total no. in group:

10

Clinical observations:

positive reactions consisted of discrete/patchy to moderate/confluent erythema

Any other information on results incl. tables

Details on skin reactions after the challenge procedure

Reading time point	24 h			48 h
	positive	total	% positive of total	positive	total	% positive total
Control group (induced without test article)
challenged with the test item 50 % in PEG 400	0	5	0	0	5	0
challenged with PEG 400	0	5	0	0	5	0
Test group (induced with test article)
challenged with the test item 50 % in PEG 400	7	10	70	7	10	70
challenged with PEG 400	0	10	0	0	10	0

Remark:

No toxic symptoms and no mortality were observed in the animals.

Applicant's summary and conclusion

Interpretation of results:

Category 1B (indication of skin sensitising potential) based on GHS criteria