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Diss Factsheets

Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Aug 1989
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1990
Report date:
1990

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
1992
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Version / remarks:
2003
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
1996
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Currently no LLNA study is available for assessment. The GPMT (depending on the test) test has been carried out as an animal test to predict human sensitisation for over a decade and is recommended by international test guidelines such as OECD.”

Test material

Constituent 1
Chemical structure
Reference substance name:
Diquat dibromide
EC Number:
201-579-4
EC Name:
Diquat dibromide
Cas Number:
85-00-7
Molecular formula:
C12H12N2.2Br
IUPAC Name:
1,1'-ethylene 2,2'-bipyridyldiylium dibromide
Constituent 2
Chemical structure
Reference substance name:
Water
EC Number:
231-791-2
EC Name:
Water
Cas Number:
7732-18-5
Molecular formula:
H2O
IUPAC Name:
water
Test material form:
liquid

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Remarks:
Alpk
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: 5 - 7 weeks (males); 7 - 10 weeks (females)
- Weight at study initiation: 278 - 418 g (males); 303 - 413 g (females)
- Housing: Individually in suspended cages (37 cm x 32 cm x 20 cm)
- Diet: Guinea Pig Diet ad libitum
- Water: Water ad libitum
- Acclimation period: Minimum of 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 ± 2
- Humidity (%): 50 ±10
- Air changes (per hr): 20 - 30
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From Aug 1989 To: Aug 1989

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
other: deionised water
Concentration / amount:
Injection: 0.05 to 0.1 mL
Treatment: Freund’s complete adjuvant (FCA) plus deionised water in the ratio 1:1
Control: Freund’s complete adjuvant (FCA) plus deionised water in the ratio 1:1
Day(s)/duration:
Day 0: First injection of the three pairs of intradermal injections
Route:
intradermal
Vehicle:
other: deionised water
Concentration / amount:
IInjection: 0.05 to 0.1 mL
Treatment: 0.1 % (w/v) preparation of the test sample in deionised water
Control: deionised water
Day(s)/duration:
Day 0: Second injection of the three pairs of intradermal injections
Adequacy of induction:
other: Well tolerated locally and systemically
Route:
intradermal
Vehicle:
other: deionised water
Concentration / amount:
Injection: 0.05 mL to 0.1 mL
Treatment: 0.1 % (w/v) preparation of the test sample in a 1:1 preparation of FCA plus deionised water
Control: a 1:1 preparation of FCA plus deionised water
Day(s)/duration:
Day 0: Third injection of the three pairs of intradermal injections
Adequacy of induction:
other: Well tolerated locally and systemically
Route:
epicutaneous, occlusive
Vehicle:
other: deionised water
Concentration / amount:
Volume/area: 0.2 - 0.3 mL/4x2 cm over the site of injection
Treatment: undiluted test substance, occlusive
Day(s)/duration:
Day 7: all tested animals for 48 hours
Adequacy of induction:
highest technically applicable concentration used
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
Volume/area: 0.2 - 0.3 mL/4x2 cm over the site of injection
Treatment: nothing applied
Day(s)/duration:
Day 7: all contol animals
Challenge
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
other: deionised water
Concentration / amount:
Volume/area: 0.05 - 0.1 mL / 1 cm x 1.5-2.0 cm area on the shorn flanks of each guinea pig.
Treatment: (1) undiluted test substance, (2) 10 % (w/v) preparation of test substance in deionised water, (3 and 4) deionised water (x2)
Day(s)/duration:
Day 21: occlusive dressing for 24 h
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
Main study: 20 test and 20 control females
Details on study design:
ANIMAL ASSIGNMENT AND TREATMENT:
The sensitisation potential of test substance was assessed using a method based on the maximisation test. Two main procedures were involved; (a) the induction of an immune response; (b) a challenge of that response.

POSITIVE CONTROLS:
A positive control study was conducted using essentially the same methodology and using formaldehyde as the test substance. The method used an intradermal induction of a 0.3 % w/v preparation in deionised water and 30 % w/v preparations in deionised water for the topical induction and challenge phase.
Challenge controls:
0.05 - 0.1 mL of deionised water
Positive control substance(s):
yes
Remarks:
formaldehyde was used as the test substance

Results and discussion

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
undiluted test substance
No. with + reactions:
4
Total no. in group:
16
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
10 % w/v preparation
No. with + reactions:
1
Total no. in group:
16
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
0 (deionised water)
No. with + reactions:
0
Total no. in group:
16
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
undiluted test substance
No. with + reactions:
0
Total no. in group:
17
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
10 % w/v preparation
No. with + reactions:
0
Total no. in group:
17
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0 (deionised water)
No. with + reactions:
0
Total no. in group:
17
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
30 % w/v aqueous dilution
No. with + reactions:
17
Total no. in group:
18
Clinical observations:
mild redness or moderate diffuse redness
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
undiluted test substance
No. with + reactions:
2
Total no. in group:
16
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
10 % w/v preparation
No. with + reactions:
0
Total no. in group:
16
Remarks on result:
no indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
0
No. with + reactions:
0
Total no. in group:
16
Remarks on result:
no indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
30 % w/v aqueous dilution
No. with + reactions:
17
Total no. in group:
18
Clinical observations:
scattered mild redness or moderate diffuse redness

Any other information on results incl. tables

MORTALITY / CLINICAL OBSERVATIONS

Two test and two control animals died, and a further test animal was killed in extremis, prior to challenge but these deaths were considered not to be treatment-related. The bandage used at challenge slipped on one test and one control animal and these animals were therefore excluded from analysis of the results.

 

BODY WEIGHTS:

There were no treatment-related effects on body weight during the study.

CHALLENGE REACTIONS AND DURATION: 

Following challenge with the undiluted test sample, scattered mild redness was seen in 5/16 test and 0/17 control animals. The net response was calculated to be 31 %.

Following challenge with a 10 % w/v preparation of the test sample in deionised water, scattered mild redness was seen in 1/16 test and 0/17 control animals. The net response was calculated to be 6 %.

No erythematous reactions were seen in any animal, test or control, following challenge with deionised water. The net response was zero.

 

POSITIVE CONTROL:

In the positive control study following challenge with a 30 % w/v preparation of a 40 % w/v aqueous formaldehyde solution, scattered mild redness or moderate diffuse redness was seen in 17/18 test and 0/10 control animals. The net response was calculated to be 94 % and formaldehyde was considered to elicit an extreme sensitisation response in previously induced guinea pigs thereby confirming the sensitivity of the test.

Table 1: Maximisation test: Number of animals with signs of allergic skin reactions

Scored after:

24 hours

48 hours

Test group

undiluted test substance

4/16

2/16

 

10 % w/v preparation

1/16

0/16

 

deionised water

0/16

0/16

 

deionised water

0/16

0/16

Vehicle Control group

undiluted test substance

0/17

0/17

 

10 % w/v preparation

0/17

0/17

 

deionised water

0/17

0/17

 

deionised water

0/17

0/17

Positive control (40 % w/v formaldehyde)

30 % w/v aqueous dilution

17/18 (0/10 control)

17/18 (0/10 control)

Applicant's summary and conclusion

Interpretation of results:
Category 1 (skin sensitising) based on GHS criteria
Conclusions:
Challenge of previously induced guinea pigs with undiluted test substance elicited a moderate sensitisation response and challenge with a 10 % w/v preparation elicited a weak sensitisation response.
Executive summary:

The sensitisation potential of the test substance was assessed in accordance with OECD 406 and in compliance with GLP using the maximisation test of Magnusson and Kligman (1970). Groups of 20 test and 20 control young adult female Alpk:Dunkin Hartley guinea pigs were used for the main study. Two main procedures were involved; (a) the induction of an immune response; (b) a challenge of that response. For the main study, the concentrations used were 0.1 % (w/v) in deionised water for the induction intradermal injections, 100 % for the topical induction applications and 10 % (w/v) in deionised water and 100 % for the challenge applications. A positive control study was conducted using essentially the same methodology and using formaldehyde as the test substance. The method used an intradermal induction of a 0.3 % w/v preparation in deionised water and 30 % w/v preparations in deionised water for the topical induction and challenge phase.


Following challenge with the test substance in form of an aqueous technical concentrate, scattered mild redness was seen in 4/16 test and 0/17 control animals. The net response was calculated to be 31 %. Following challenge with a 10 % w/v preparation of the test sample in deionised water, scattered mild redness was seen in 1/16 test and 0/17 control animals. The net response was calculated to be 6 %. No erythematous reactions were seen in any test or control animal following challenge with deionised water. The net response was zero. In the positive control study following challenge with a 30 % w/v preparation of a 40 % w/v aqueous formaldehyde solution, scattered mild redness or moderate diffuse redness was seen in 17/18 test and 0/10 control animals. The net response was calculated to be 94 % and formaldehyde was considered to elicit an extreme sensitisation response in previously induced guinea pigs thereby confirming the sensitivity of the test.


Challenge of previously induced guinea pigs with undiluted test substance elicited a moderate sensitisation response and challenge with a 10 % w/v preparation elicited a weak sensitisation response.