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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1988
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1988
Report Date:
1988

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
adopted 12 May 1981
GLP compliance:
yes
Type of study:
guinea pig maximisation test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Nitro-m-dichlorbenzol 413 (2,4-Dichloro-1-nitrobenzene)
- Physical state: yellow crystals
- Analytical purity: > 98 %
- Impurities (identity and concentrations): Dichloronitrobenzene-isomeres
- Storage condition of test material: 4°C, in the dark

In vivo test system

Test animals

Species:
guinea pig
Strain:
other: Pirbright-White
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Hoechst AG, Kastengrund, SPF-Zucht
- Weight at study initiation: 326 +/- 30 g
- Housing: 5 animals per cage (Macrolon type 4)
- Diet (e.g. ad libitum): ERKA-Mischfutter Nr. 8300 für Meerschweinchen und Kaninchen, ad libitum
- Water (e.g. ad libitum): tap-water ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 50 +/- 20
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
other: paraffin oil and petrolatum
Concentration / amount:
Intradermal induction: 5% in paraffin oil
Epicutaneous induction: 25% in petrolatum
Epicutaneous challenge: 25% in petrolatum
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
other: paraffin oil and petrolatum
Concentration / amount:
Intradermal induction: 5% in paraffin oil
Epicutaneous induction: 25% in petrolatum
Epicutaneous challenge: 25% in petrolatum
No. of animals per dose:
10
Details on study design:
RANGE FINDING TESTS:
- Determination of the primary not irritating concentration:
The following conentrations of the test substance were tested (epicutaneous, occlusive) on the left flank of two guinea pigs: 0.25, 2.50 and 25.00 % in petrolatum.
The left flank of the test animals was clipped, 0.5 g of the test substance preparation was loaded onto a 2x2 cm cellulose layer, which was occlusively fixed to the left flank for 24 h. The treated skin was observed 24 h after removing the patch and evaluated for erythema/edema according to Draize score.
- Determination of the tolerance for intradermal injection:
The following test substance conentrations were tested twice in 3 guinea pigs by intradermal injection: 0.2, 1.0 and 5.0 % in paraffin oil.
The injections sites were localized within an area of 2x4 cm on the back of the animals near the shoulders.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 (intradermal and epicutaneous)
- Exposure period:
intradermal: 8 days
epicutaneous: 48 h
- Test groups:
intradermal: 50% Freund's adjuvant, 5% test substance in paraffin oil, 5% test substance in Freund's adjuvant
epicutaneous: 25% test substance in petrolatum
- Control group:
intradermal: 50% Freund's adjuvant, paraffin oil, Freund's adjuvant
epicutaneous: petrolatum
- Site:
intradermal: The injections sites were localized within an area of 2x4 cm on the back of the animals near the shoulders.
epicutaneous: The test substance was applied on the same area for the intradermal injections.
- Frequency of applications:
intradermal: 2
epicutaneous: 1


B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 1
- Exposure period: 24 h
- Test groups: The left flank of the test animals was clipped (5x5 cm). 0.5 g of the test substance preparation (25% test substance in petrolatum) was loaded onto a 2x2 cm cellulose layer, which was occlusively fixed to the left flank for 24 h.
- Control group: The left flank of the test animals was clipped (5x5 cm). 0.5 g of the test substance preparation (25% test substance in petrolatum) was loaded onto a 2x2 cm cellulose layer, which was occlusively fixed to the left flank for 24 h.
- Evaluation (hr after challenge): 48 and 72 h


Results and discussion

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
48
Group:
negative control
Dose level:
25% in petrolatum
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 25% in petrolatum. No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
25% in petrolatum
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
All animals showed very slight to moderate erythema and very slight to slight edema. In addition, some animals (3/10) showed skin drying and cracking.
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 48.0. Group: test group. Dose level: 25% in petrolatum. No with. + reactions: 10.0. Total no. in groups: 10.0. Clinical observations: All animals showed very slight to moderate erythema and very slight to slight edema. In addition, some animals (3/10) showed skin drying and cracking..
Reading:
2nd reading
Hours after challenge:
72
Group:
negative control
Dose level:
25% in petrolatum
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 72.0. Group: negative control. Dose level: 25% in petrolatum. No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
2nd reading
Hours after challenge:
72
Group:
test group
Dose level:
25% in petrolatum
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
All animals showed very slight to moderate erythema and very slight to slight edema. In addition, some animals (6/10) showed skin drying and cracking.
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 72.0. Group: test group. Dose level: 25% in petrolatum. No with. + reactions: 10.0. Total no. in groups: 10.0. Clinical observations: All animals showed very slight to moderate erythema and very slight to slight edema. In addition, some animals (6/10) showed skin drying and cracking..

Any other information on results incl. tables

Determination of the primary not irritating concentration:

After treatment with 0.25, 2.50 and 25.00% test substance preparation no signs of skin irritation were observed.

Treating the animals with Freund's adjuvant may decrease the limit concentration determined for primary irritation. For this reason, 5 Freund's adjuvant-treated animals of a satellite group were treated with 25% test substance in petrolatum by epicutaneous application. These animals showed no signs of irritation. Therefore, a concentration of 25% test substance in petrolatum was established for the challenge exposure in the main experiment.

Determination of the tolerance for intradermal injection:

The injection of 0.2, 1.0 ad 5.0% test substance preparations induced skin redness and swelling. On the basis of this preliminary test, the 5% test substance preparation was chosen for intradermal injection in the main experiment.

Main experiment

- Body weight and clinical observations:

Treated animals showed no signs of toxicity during the whole test period.

Intradermal injection with Freund's adjuvant (with and without test substance) clearly induced skin redness and swelling at the injection site. From the third day post-application, the injection sites were hardened. After 5 days post-application, necrosis was observed in most of the Freund's adjuvant-treated injection sites.

After removal of the occlusive bandage on day 11, the application sites which had been treated with Freund's adjuvant showed skin redness, swelling and were hardened in all animals (control, test and satellite groups). In addition, necrosis and some open wounds were observed. The injection sites treated only with the test substance or vehicle showed no irritation.

The body weight development of the treated animals was not affected.

- Challenge exposure:

24 and 48 h after removal of the occlusive bandage, all animals of the test group showed very slight to moderate erythema and very slight to slight edema. In addition, some animals showed skin drying and cracking. Animals of the control group showed no irritation.

48 h post-application

Control animals (No.)

1

2

3

4

5

 

 

 

 

 

Erythema score

0

0

0

0

0

 

 

 

 

 

Edema score

0

0

0

0

0

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Test animals (No.)

11

12

13

14

15

16

17

18

19

20

Erythema score

1

3

3

2

2

2

1

1

2

1

Edema score

1

2

1

1

2

1

1

1

1

1

Dry, cracked skin

 

 

 

x

 

x

 

 

x

 

 

 

 

 

 

 

 

 

 

 

 

 

72 h post-application

Control animals (No.)

1

2

3

4

5

 

 

 

 

 

Erythema score

0

0

0

0

0

 

 

 

 

 

Edema score

0

0

0

0

0

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Test animals (No.)

11

12

13

14

15

16

17

18

19

20

Erythema score

1

3

3

2

2

2

1

1

2

1

Edema score

1

2

2

1

1

1

1

0

1

1

Dry, cracked skin

 

 

 

x

 

x

x

x

x

x

Applicant's summary and conclusion

Interpretation of results:
sensitising
Remarks:
Migrated information