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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:
1991
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Well documented, according to accepted guidelines

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report Date:
1991

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
reliability scoring based on 2001 guideline
Deviations:
no
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): 1,4-Butanedioldiglycidylether (BDDGE)
- Physical state: liquid
- Analytical purity: 97.1%
- Lot/batch No.: DC 1294.2
- Storage condition of test material: room temperature in the dark

In vivo test system

Test animals

Species:
guinea pig
Strain:
other: Ibm: GOHI SPF
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: BRL, Biological Research Laboratories Ltd.
- Age at study initiation: 7 to 8 weeks
- Weight at study initiation: 314 to 346 g
- Housing: individually in Makrolon type-3 cages
- Diet (e.g. ad libitum): pelleted standard ad libitum
- Water (e.g. ad libitum): tap water ad libitum
- Acclimation period: one week


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

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
other: bi-distilled water
Concentration / amount:
1) Induction:
Intradermal injection: 1% (with bi-distilled water) of 0.1 mL
Epidermal application: 10% (in bi-distilled water) on a saturated 2 x 4 cm patch of filter paper
2) Challenge: 5% in bi-distilled water
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
other: bi-distilled water
Concentration / amount:
1) Induction:
Intradermal injection: 1% (with bi-distilled water) of 0.1 mL
Epidermal application: 10% (in bi-distilled water) on a saturated 2 x 4 cm patch of filter paper
2) Challenge: 5% in bi-distilled water
No. of animals per dose:
Range finding tests: 1 male and 1 female for the intracutaneous test and 6 males and 5 females for the epicutaneous tests.
Main study: 5 animals/sex for control group and 10 animals/sex for the test group
Details on study design:
RANGE FINDING TESTS: The objective of preliminary study was to identify irritation test article concentrations suitable for the induction phase of the main study. In addition, a suitable non-irritant concentration of the test article, by the topical route of administration, was identified for the challenge application.
1) Intradermal injections: Intradermal injections (0.1 mL/site) were made into the clipped flank of two guinea-pigs at concentrations of 5, 3, and 1% of the test article in bi-distilled water. The resulting dermal reactions were assessed 24 hours later.
2) Epidermal application: Patches of filter paper (2 x 2 cm) were saturated with concentrations of 25, 15, 10, and 5% of the test article in bi-distilled water and applied to the clipped and shaved flanks of each of four guinea-pigs. The patches were covered by a strip of aluminum foil and firmly secured by elastic plaster wrapped around the trunk and covered with impervious adhesive tape. The dressings were removed after an exposure period of 24 hours and the reaction sites were assessed for erythema and edema on a numerical basis according to the scale 24 and 48 hours after removal of the dressings. Two previous epidermal pre-tests were performed as described above, one with the undiluted test article and 75, 50, and 25% test article in vaseline, the second with the same concentrations in bi-distilled water. This latter was performed because the test article was not miscible in vasilne. The third epidermal pre-test described above was performed to confirm the previous results and to determine the slightly irritating and highest non-irritating concentration.


MAIN STUDY
A. INDUCTION EXPOSURE
I) Intradermal Injections:
An area of dorsal skin from the scapular region was clipped free of hair. Three pairs of intradermal injections (0.1mL/site) were made at the border of a 4 x 6 cm area in the clipped region as follows:
-Test group:
1) Freund's complete adjuvant 50:50 with bi-distilled water;
2) The test article, diluted to 1% with bi-distilled water; and
3) The test article diluted to 1% with bi-distilled water, emulsified in a 50:50 mixture of Freund's complete adjuvant and bi-distilled water.
-Control group:
1) Freund's complete adjuvant 50:50 with bi-distilled water;
2) Bi-distilled water; and
3) Freund's complete adjuvant 50:50 with bi-distilled water.
II) Epidermal Applications:
-Test group:
One week after the injections, the scapular area was again clipped and shaved free of hair. A 2 x 4 cm patch of filter paper was saturated with the test article (10% in bi-distilled water) and placed over the injection sites of the test animals. The patch was covered by aluminum foil and firmly secured by an elastic plaster wrapped around the trunk of the animal and secured with impervious adhesive tape. The dressings were left in place for approximately 48 hours. Reaction sites were assessed for erythema and edema 24 and 48 hours after removal of the dressing, using the numerical grading system.
-Control group:
The guinea-pigs of the control group were treated as described above with the omission of test article.

B. CHALLENGE EXPOSURE
The test and control guinea-pigs were challenged 2 weeks after the epidermal induction application. Hair was clipped and shaved from a 5 x 5 cm area on the left and right flank of each guinea-pig.
-Test group:
Two patches of filter paper were saturated with:
a) non-irritant concentration (5% in bi-distilled water) of the test article and
b) with the vehicle only
and applied to the left flank and right flank using the same method as for the epidermal application.
The dressings were removed approximately 24 hours later. The sites were assessed for erythema and edema 24 and 48 hours after removal of the dressing, using the numerical scoring system as described under preliminary study. Erythema and edema reactions were described. The challenge site was evaluated 24 and 48 hours after removal of the patch. The readings were made under artificial fluorescent light (daylight spectrum). The reactions were scored on the basis of the Draize score. Based upon the percentage of animals sensitised (24-hour reading), the test article was assigned to one of the following 5 grades of allergenic potency, ranging from weak to extreme.
Sensitisation rate (%): Grade: Classification
0-8: 1: Weak
9-28: 2: Mild
29-64: 3: Moderate
65-80: 4: Strong
81-100: 5: Extreme
-Control group:
The control animals were treated in the same way as described above.

OTHER: Erythema and edema were assessed using the following numerical grading system according to Draize:

Erythema and eschar formation:
No erythema:0
Very slight erythema (barely perceptible): 1
Well-defined erythema: 2
Moderate to severe erythema: 3
Severe erythema (beet redness) to slight eschar formation (injuries in depth): 4

Edema formation:
No edema: 0
Very slight edema (barely perceptible): 1
Slight edema (edges of area well-defined by definite raising): 2
Moderate edema (raised approximately 1 mm): 3
Severe edema (raised more than 1 mm and extending beyond the area of exposure): 4
Challenge controls:
See details on study design.
Positive control substance(s):
yes
Remarks:
formaldehydesung

Study design: in vivo (LLNA)

Vehicle:
other: not applicable
Concentration:
not applicable
No. of animals per dose:
not applicable
Details on study design:
not applicable
Positive control substance(s):
other: not applicable
Statistics:
Irritation graded according to scale. Mean values were reported with standard deviations. Fisher-test was used and for calculation of p-values the 24-hour reading of the animals from the control and test group was used.

Results and discussion

Positive control results:
For the induction period, a 20% dilution of formaldehydesung (HCHO) in bi-distilled water and for the challenge procedure, a 15% dilution of HCHO was used. Clear positive results were observed in the HCHO-treated animals after the epidermal challenge application (7 positive reactions among 10 animals tested). According to the results observed, it is considered that HCHO possess an strong skin sensitising (contact allergenic) potential in the guinea pig strain used (Ibm: GOHI; SPF-quality).

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
5%
No. with + reactions:
2
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: 5%. No with. + reactions: 2.0. Total no. in groups: 10.0. Clinical observations: none.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
5%
No. with + reactions:
1
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: 5%. No with. + reactions: 1.0. Total no. in groups: 10.0. Clinical observations: none.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
5%
No. with + reactions:
17
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: 5%. No with. + reactions: 17.0. Total no. in groups: 20.0. Clinical observations: none.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
5%
No. with + reactions:
15
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: 5% . No with. + reactions: 15.0. Total no. in groups: 20.0. Clinical observations: none.
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
15%
No. with + reactions:
7
Total no. in group:
10
Clinical observations:
not reported
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: positive control. Dose level: 15%. No with. + reactions: 7.0. Total no. in groups: 10.0. Clinical observations: not reported.
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
15%
No. with + reactions:
3
Total no. in group:
10
Clinical observations:
not reported
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: positive control. Dose level: 15%. No with. + reactions: 3.0. Total no. in groups: 10.0. Clinical observations: not reported.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: not applicable
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: not applicable

Any other information on results incl. tables

Sensitizing Effects

CONTROL GROUP:

Two out of 10 and 1 out of 10 animals showed spontaneous positive erythema reactions after the first challenge when treated with the 5% test article dilution at the 24- and 49- hour reading, respectively. No positive erythema reactions were observed with bi-distilled water alone.

 

TEST GROUP:

Seventeen out of 20 and 15 out of 20 animals showed positive erythema reactions after the first challenge when treated with the 5% test article dilution at the 24- and 48- hour reading, respectively. No positive erythema reactions were observed with bi-distilled water alone.

 

MORTALITY/VIABILITY

No death occurred during the study.

 

SYMPTOMS, LOCAL

CONTROL GROUP:

Application area around the injection sites 1and 3 was found to show erythema and edema from day 2 to 7; necrosis were observed from day 8 to 21 and encrustations from day 20 to 31. No local symptoms were observed in injection site 2.

 

TEST GROUP:

Application area around the injections sites 1, 2 and 3 was found to show the same local symptoms as described above for the injections sites1 and 3 of control group.

 

On day 9 of test no observation could be performed because the animals were bandaged semi-occlusively.

 

SYMPTOMS, SYSTEMIC

No systemic symptoms were observed in the animals.

 

BODY WEIGHTS

The body weight gain of the animals was not affected adversely during the study.

Applicant's summary and conclusion

Interpretation of results:
sensitising
Remarks:
Migrated information