Disodium 3,3'-dithiobis[propanesulphonate]

Endpoint summary

Description of key information

Skin sensitisation: Guinea pig maximisation test, Guinea pig (Dunkin-Hartley), male, SPS, OECD 406, EU method B.6, GLP: sensitising
Skin sensitisation: Bühler test, Guinea pig (Dunkin-Hartley), MPS, OECD 406, GLP: not sensitising

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:

16 June - 18 July 1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP - Guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

(The Department of Health of the Government of the United Kingdom)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

No new study has been conducted; data from a existing GLP-guideline study (1997) has been used as this study is totally sufficient to cover this endpoint.

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: David Hall Limited , Burton-on-Trent , Staffordshire , UK
- Age at study initiation: approximately 8 - 12 weeks
- Weight at study initiation: 310 - 406 g
- Housing: animals were housed singly or in pairs in solid-floor polypropylene cages furnished with woodflakes .
- Diet : Guinea Pig FD1 Diet (Special Diets Services Limited , Witham , Essex , UK) , ad libitum
- Water : tap water , ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 . 23
- Humidity (%): 52 - 76
- Air changes (per hr): approximately 15
- Photoperiod (hrs dark / hrs light): 12/12

Route:

intradermal and epicutaneous

Vehicle:

water

Concentration / amount:

Intradermal Induction : 10%
Topical Induction : 50%
Topical Challenge : 50% and 25%

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

Intradermal Induction : 10%
Topical Induction : 50%
Topical Challenge : 50% and 25%

No. of animals per dose:

10 in test group , 5 in control group

Details on study design:

RANGE FINDING TESTS:
Selection of concentration for Intradermal Induction : Intradermal injections were made on 4 guinea pigs using concentrations of 1 ,5 ,10 and 25% w/v in distilled water . The degree of erythema at the injection sites was assessed approximately 24 , 48 and 72 hours and 7 days after injection . The degree of oedema was not evaluated . Any evidence of systemic toxicity was also recorded . The highest concentration that caused only mild to moderate skin irritation , and which was well tolerated systemically , was selected for the intradermal induction stage of the main study .

Selection of Concentration for Topical Induction : Two guinea pigs (intradermally injected with Freund´s Complete Adjuvant eight days earlier) were treated with four preparations of the test material (50% , 25% , 10% and 5% w/w in distilled water) . Applications were made to the clipped flanks under occlusive dressings for an exposure period of 48 hours . The degree of erythema and oedema was evaluated approximately 1 , 24 and 48 hours after dressing removal . The highest concentration producing only mild to moderate dermal irritation was selected for the topical induction stage of the main study .

Selection of Concentration for Topical Challenge : Four preparations of the test material (50% , 25% , 10% and 5% w/w in distilled water) were applied to the clipped flanks of two guinea pigs under occlusive dressings for an exposure period of 24 hours . These guinea pigs did not form part of the main study but had been treated identically to the control animals of the main study , up to Day 14 . The degree of erythema and oedema was evaluated approximately 1 , 24 and 48 hours after dressing removal . The highest non-irritant concentration of the test material and one lower concentration were selected for the topical challenge stage of the main study .

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 , intradermal and epicutaneous
- Exposure period: single injection (intradermal , left and right side) and 48 hours (epicutaneous)
- Test groups:

Intradermal (3 pairs of injections , 0.1 ml) :
Injection 1 : a 1:1 mixture (v/v) FCA/distilled water
Injection 2 : a 10% (w/v) solution of the test material in distilled water
Injection 3 : a 10% (w/v) emulsion of the test material in a 1:1 preparation of FCA/distilled water
24 and 48 h after intradermal injection , the degree of erythema was evaluated .

Epicutaneous :
50% (w/w) test material in distilled water
On day 7 the animals were treated with a topical application of the test material . The occlusive dressing was kept in place for 48 h . The degree of erythema and oedema was evaluated 1 and 24 h after removing the patch .

-Control Group :
Intradermal (3 pairs of injections) :
Injection 1 : a 1:1 mixture (v/v) FCA/distilled water
Injection 2 : distilled water
Injection 3 : a 50% (w/v) formulation of distilled water in FCA/distilled water 1:1
Epicutaneous : distilled water

- Site : approximately 40 mm x 60 mm area on the shoulder region
- Frequency of applications : once (intradermal and epicutaneous)
- Duration : day 0 (intradermal) , day 7 - 9 (epicutaneous)
- Concentrations : 10% (intradermal) , 50% (epicutaneous)


B. CHALLENGE EXPOSURE
- No. of exposures: one , topical
- Day(s) of challenge: 21
- Exposure period: 24 hours
- Test groups: test material in distilled water
- Control group: test material in distilled water
- Site: approximately 50 mm x 70 mm , on both flanks of the animals
- Concentrations: 50% on the right flank , 25% on the left flank
- Evaluation (hr after challenge): 24 and 48 hours after exposure ended

OTHER: remaining test material was removed from the test site after epicutaneous exposure lasting 24 h .

Positive control substance(s):

yes

Remarks:

(historical data using 2,4-Dinitrochlorobenzene , Neomycin Sulphate and 2-Mercaptobenzothiazole as positive controls was included)

Positive control results:

Historical data on three positive control substances show that they induced positive reactions in control animals , thus meeting the reliability criteria for the GPMT .
In three independent studies performed in 1994 , 1995 and 1996 2,4-Dinitrochlorobenzene induced sensitation in 100% of the animals. A 0.1% solution in arachis oil was used for intradermal induction , 1% solution in arachis oil and 0.75% solution in ethanol for topical induction (0.05% and 1% in arachis oil , 0.1% and 0.25% in ethanol for topical challenge)
In one study performed in 1995 Neomycin Sulphate induced sensitation in 60% of the animals. A 10% solution in distilled water was used for intradermal induction , 75% solution in distilled water for topical induction , and 50% and 75% in distilled water for topical challenge.
In two independent studies performed in 1996 2-Mercaptobenzothiazole induced sensitation in 70-90% of the animals. A 10% solution in arachis oil was used for intradermal induction , 50% solution in acetone:PEG400 (70:30) for topical induction , and 25% and 50% in acetone:PEG400 (70:30) for topical challenge.

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

25%

No. with + reactions:

9

Total no. in group:

10

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 25%. No with. + reactions: 9.0. Total no. in groups: 10.0.

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

50%

No. with + reactions:

10

Total no. in group:

10

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 50%. No with. + reactions: 10.0. Total no. in groups: 10.0.

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

25%

No. with + reactions:

1

Total no. in group:

5

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 25%. No with. + reactions: 1.0. Total no. in groups: 5.0.

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

50%

No. with + reactions:

1

Total no. in group:

5

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 50%. No with. + reactions: 1.0. Total no. in groups: 5.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

25%

No. with + reactions:

9

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 25%. No with. + reactions: 9.0. Total no. in groups: 10.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

50%

No. with + reactions:

10

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 50%. No with. + reactions: 10.0. Total no. in groups: 10.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

25%

No. with + reactions:

1

Total no. in group:

5

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 25%. No with. + reactions: 1.0. Total no. in groups: 5.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

50%

No. with + reactions:

1

Total no. in group:

5

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 50%. No with. + reactions: 1.0. Total no. in groups: 5.0.

Group:

positive control

Dose level:

concurrent control data, various substances and dose levels

Remarks on result:

positive indication of skin sensitisation

Remarks:

% incidence of sensitization ranged in several studies from 60% to 100%

Bodyweight gains of guinea pigs in the test group , between Day 0 and Day 24 were comparable to those observed in the control group animals over the same period .

Very slight to well-defined erythema was noted at the intradermal induction sites of all test group animals at the 24-hour observation . Very slight to well-defined erythema was noted at the intradermal induction sites of five test group animals at the 48-hour observation .

Very slight to well-defined erythema was noted at the intradermal induction sites of all control group animals at the 24-hour observation . Very slight to well-defined erythema was noted at the intradermal induction sites of two control group animals at the 48-hour observation .

Very slight to well-defined erythema and very slight to slight oedema were noted at the topical induction sites of all test group animals at the 1-hour observation .Very slight erythema with or without very slight oedema was noted at the topical induction sites of nine test group animals at the 24-hour observation . Small superficial scattered scabs were noted at the induction sites of five test group animals at the 24-hour observation . Bleeding from the injection sites was noted in 4 treated and 3 control group animals .

Topical Challenge :

50% w/w in distilled water

Positive skin responses (moderate to severe erythema - grade 3 and slight oedema) were noted at two test group animals at the 24 -hour observation and persisted in one test group animal at the 48-hour observation . Very slight to well-defined erythema with or without very slight to slight oedema was noted at eight test group animals at the 24 -hour observation and in nine test group animals at the 48-hour observation . Small superficial scattered scabs were noted at the challenge sites of two test group animals at the 24 and/or 48-hour observations . The reaction extended beyond the challenge site of one test group animal at the 24 and 48-hour observations .

Well-defined erythema and very slight oedema were noted at one control group animal at the 24-hour observation with very slight erythema at the 48-hour observation

25% w/w in distilled water

Positive skin responses (well-defined or moderate to severe erythema - grades 2 or 3 and very slight to slight oedema) were noted at nine test group animals at the 24-hour observation and in eight test group animals at the 48-hour observation . Very slight erythema and very slight oedema were noted at one test group animal at the 48-hour observation . Small superficial scattered scabs were noted at the challenge sites of two test group animals at the 48-hour observations .

Very slight erythema was noted at one control group animal at the 24 and 48-hour observations.

Table 1 : Intradermal induction , individual skin reactions

	Grade of Erythema at Observation Time
	24 Hours	24 Hours	48 Hours	48 Hours
Animal No.	Left Side	Right Side	Left Side	Right Side
1 test group	2	2	0	1
2 test group	2	2	0	0
3 test group	2	1	1	0
4 test group	1	0	1	0
5 test group	1	1	1	1
6 test group	2	2	0	0
7 test group	2	2	0	0
8 test group	2	2	0	0
9 test group	1	1	0	0
10 test group	2	1	1	0
11 control group	1	0	0	0
12 control group	1	1	0	0
13 control group	2	2	1	1
14 control group	1	0	1	0
15 control group	1	1	0	0

Interpretation of results:

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

Conclusions:

The study was performed according to the OECD TG406 without deviations and therefore considered to be of the highest quality (reliability Klimisch 1). The validity criteria of the test system are fulfilled. The test material produced a 90% (9/10) sensitisation rate and was classified as an extreme sensitiser to guinea pig skin . The test material was also classified as a sensitiser according to EU labelling regulations .

Executive summary:

A study was performed to assess the contact sensitisation potential of the test material in the albino guinea pig. The study was performed in compliance with the OECD Guidelines for Testing of Chemicals No. 406 "Skin Sensitisation (adopted 17 July 1992) and Method B6 of Commission Directive 92/69/EEC (which constitutes Annex V of Council Directive 67/548/EEC). Ten test and five control animals were used for the main study.

Based on the results of sighting tests, the concentrations of test material for the induction and challenge phases were selected as follows:

-Intradermal Induction 10% w/v in distilled water

-Topical Induction 50% w/w in distilled water

-Topical Challenge 50% and 25% w/w in distilled water

The test material produced a 90% (9/10) sensitisation rate and was classified as an extreme sensitiser to guinea pig skin. The test material was also classified as a sensitiser according to EU labelling regulations. The Symbol "Xi", the indication of danger 'irritant' and the risk phrase R 43 "MAY CAUSE SENSITISATION BY SKIN CONTACT" are required.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

There is an in vivo Klimisch 1 GLP OECD 406 guideline study on SPS itself available. It was performed as a maximisation test. This testing procedure aims to detect all possible sensitizing effects by including intradermal injections during the induction phase. This scenario is not totally realistic for human exposure and possible overestimates a sensitising potential of a substance. In the available study, the test material produced a 90% (9/10) sensitisation rate and was classified as an extreme sensitiser to guinea pig skin. The test material was also classified as a sensitiser according to EU labelling regulations (Directive 67/548/EEC).

The supporting GLP OECD 406 guideline study on MPS was performed using the method of Bühler. The Bühler assay only employs epicutaneous induction, which is more related to real human exposure. In this study, the test material did not induce any response during the 3 weeks of induction treatment and during the challenge phase on the skin of guinea pigs. Therefore the substance is not considered to be a skin sensitizer under conditions which are more relavant for human exposure. SPS and MPS are considered comparable as data indicated they are transformed in vivo into each other, hence bearing the same protein-binding potential which is considered relevant for any response of the immune system.

However, as a precautionary and conservative approach, it is considered reasonable to classify SPS as skin sensitizer as a risk of sensitisation cannot completely excluded. Following this approach, no further testing is considered necessary. One may consider that the actual sensitizing effects of SPS in humans are expected to be less severe than indicated by the present data on skin sensitisation in guinea pigs and by classification.

Justification for selection of skin sensitisation endpoint:
Klimisch 1 key study on SPS itself.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

In the available key study on SPS (GPMT), which is considered most reliable, the test item was found to be an extreme sensitiser to guinea pig skin. The test material was also classified as a sensitiser according to EU labelling regulations (Directive 67/548/EEC). These findings are similarly relevant for the recent C&L Regulation (EC) 1272/2008 and do also trigger classification as a sensitiser. As an precautionary and conservative approach, SPS is classified as skin sensitizer Cat. 1B because 90% reacted to an intradermal induction with 10% SPS. The boundary values according to Regulation (EC) 1272/2008 are i.a. ≥ 30 % responding at > 1 % intradermal induction dose in the GPMT. The classification seems reasonable, although other data indicate that the actual sensitization potential under real-life conditions may be way less severe.