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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:
14 January - 7 February 1997
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1997
Report Date:
1997

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
adopted July 17, 1992
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
July 17, 1992
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
The Murine Local Lymph Node Assay (LLNA) is the first-choice method for in vivo testing according to the REACH Regulation. However, this reliable GPMT test was performed before entry into force of the REACH Regulation.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid: volatile
Details on test material:
Batch No: Tank B 612
Date of manufacturing : August 1996
Storage conditions: Room temperature (storage temperature =<25 °C), exclusion of oxygen (storage under nitrogen)

In vivo test system

Test animals

Species:
guinea pig
Strain:
Pirbright-Hartley
Remarks:
Pirbright White, Dunkin Hartley. Crl : (HA) BR [SPF]
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River GmbH - Wiga, Kisslegg, FRG
- Females (if applicable) nulliparous and non-pregnant: not specified
- Age at study initiation: Young adult animals
- Weight at study initiation: 321 - 397 g
- Housing (cage type): Makrolon, type IV, 5 animals per cage; bedding: Granulat type 3/4 (staubfrei) SSNIFF
- Diet: Kliba Labordiät 341 (Kaninchen-Meerschweinchen-Haltungsdiät) ad libitum
- Water: Water ad libitum (tap water; about 2 g of ascorbic acid per 10 L water was added to the drinking water twice a week)
- Acclimation period: 7 days before the beginning of the study
- Indication of any skin lesions: not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 – 24
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
olive oil
Concentration / amount:
0.1 mL conc: 5% in olive
Day(s)/duration:
1
Adequacy of induction:
other: based on the pretests
Route:
other: Percutaneous occlusive
Vehicle:
unchanged (no vehicle)
Day(s)/duration:
2
Adequacy of induction:
other: based on the pretests
Challenge
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
olive oil
Concentration / amount:
concentration: 75%
Day(s)/duration:
1
Adequacy of challenge:
not specified
No. of animals per dose:
Control group: 10
Test group: 20
Details on study design:
Clipping of the test animals: at least 2 hours before each test substance application at the appropriate application sites.
1) PRETEST:
Amount applied: 2 x 2 cm filter paper strips containing the test substance or test substance formulation were applied to the skin of the flanks under an occlusive dressing. The filter paper strip was soaked in the test substance or test substance formulation. The dressing consisted of rubberized linen patches (4 x 4 cm. from Russka), patches of Idealbinde (5 x 5 cm from Pfdlzische Verbandstoff-Fabrik) and Fixomull@ Stretch (adhesive fleece) from Beiersdorf A.G .
Exposure period: The test substance was applied 2 times for 24 hours within a period of 96 hours in order to detect nonspecific phenomena that are not caused by a sensitization reaction but could possibly be attributed to a shift in the irritation threshold.
Site of application: flank, on the same area respectively
Number of test animals: 4 per test concentration
Readings: about 24 and 48 h after the beginning of application

2) MAIN TEST:
A. INDUCTION EXPOSURE
Intradermal induction: 6 intradermal injections in groups of two per animal
Test group:
A) front row: 2 injections each of 0 .1 ml Freund's adjuvant* without test substance emulsified with 0 .9% aqueous NaCl-solution in a ratio of 1:1
B) middle row: 2 injections each of 0 .1 ml of the test substance formulation
C) back row: 2 injections each of 0 .1 ml Freund's adjuvant* / 0 .9% aqueous NaCl-solution (1:1) with test substance
Control group: Same injections (A, B, C) but without test substance, only with the formulating agent
Site: shoulder
Duration of exposure: 48 hours
Readings: 24 h after the beginning of application

Percutaneous induction:
Percutaneous induction was carried out one week after intradermal induction.
Amount: 2 x 4 cm. filter paper strips containing the test sub stance were applied to the skin of the shoulder under an occlusive dressing. The filter paper strip was soaked in the test substance. The dressing consisted of rubberized linen patches (4 x 6 cm from Russka) and FixomullO Stretch (adhesive fleece) from Beiersdorf AG. The control groups were not treated, since the test substance was applied undiluted and thus no solvent was used.
Site: shoulder, same area as in the case of the previous intradermal application
Duration of exposure: 48 hours
Readings: 48 h after the beginning of application

B. CHALLENGE EXPOSURE
The challenge was performed 14 days after the percutaneous induction.
Amount: 2 x 2 cm filter paper strips containing the test substance formulation were applied to the skin of the flank under an occlusive dressing. The filter paper strip was soaked in the test substance formulation. The dressing consisted of rubberized linen patches (4 x 4 cm from Russka), patches of Idealbinde (5 x 5 cm from Pfdlzische Verbandstoff-Fabrik) and Fixomull Stretch (adhesive fleece) from Beiersdorf AG. The test group and control group 1 were treated with the test substance formulation (control group 2 remained untreated).
Duration of exposure: 24h
Site: intact flank
Evaluation: 24 and 48h after removal of the patch
Challenge controls:
The number of animals with skin findings at 24 and/or 48 hours after the removal of the patch is taken into account for the determination of the sensitization rate. The evaluation "sensitizing" results if at least 30 per cent of the test animals exhibit skin reactions in this adjuvant test.
Positive control substance(s):
yes
Remarks:
Alpha-Hexylcinnamaldehyde tech. 85%

Results and discussion

Positive control results:
Positive control (reliability check) with a known sensitizer is not included in this study. However, a separate study is performed twice a year in the laboratory. The positive control with Alpha-Hexylcinnamaldehyde techn. 85% showed that the test system was able to detect sensitizing compounds under the laboratory conditions chosen.

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
75% of the test substance
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no clinical sign
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
24
Group:
test group
Dose level:
75% of the test substance
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no clinical sign
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
75% of the test substance
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
no clinical sign
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
75% of the test substance
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
no clinical sign
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
5% positive control in Lutrol E 400 DAB
No. with + reactions:
11
Total no. in group:
18
Clinical observations:
no clinical signs
Remarks on result:
positive indication of skin sensitisation
Remarks:
Very slight erythema could be observed in 6 animals. 3 animals exhibited well-defined erythema, whereas 1 out of these animals additionally showed very slight edema . In 2 animals moderate erythema and very slight edema were observed.
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
5% positive control in Lutrol E 400 DAB
No. with + reactions:
5
Total no. in group:
18
Clinical observations:
no clinical signs
Remarks on result:
positive indication of skin sensitisation
Remarks:
Very slight erythema was seen in 2 animals, whereas 1 out of these animals additionally exhibited scaling . Well-defined erythema could be observed in 2 animals. 1 out of these aditionally showed very slight edema and superficial scabbing. Moderate erythema, very slight edema and superficial scabbing was seen in 1 animal . 2 animals exhibited scaling.
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
positive control
Dose level:
5% positive control in Lutrol E 400 DAB
No. with + reactions:
8
Total no. in group:
18
Clinical observations:
no clinical signs
Remarks on result:
positive indication of skin sensitisation
Remarks:
Very slight erythema in 4 animals. Well-defined erythema in 4 animals and 1 out of these animals additionally showed scaling.
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
positive control
Dose level:
5% positive control in Lutrol E 400 DAB
No. with + reactions:
5
Total no. in group:
18
Clinical observations:
no clinical signs
Remarks on result:
positive indication of skin sensitisation
Remarks:
Very slight erythema was noted in 4 animals, 2 out of these animals additionally exhibited scaling. Well-defined erythema was noted in 1 animal .

Any other information on results incl. tables

RESULTS OF THE PRETEST:

The pretest was performed with 4 female animals for percutaneous and with 2 female animals for intradermal induction.

Two 24-hour percutaneous occlusive applications within 96 hours were performed. The minimum irritant concentration was found to be the undiluted test substance . The maximum non-irritant concentration was found to be a 75% test substance preparation in olive oil DAB 10.

Applicability: it was possible to inject a 5% test substance preparation in olive oil DAB 10 resp . in Freund's adjuvant / 0 .9% aqueous NaCl-solution (1 : 1) with a syringe.

RESULTS OF THE MAIN TEST:

- Induction:

After the intradermal induction well-defined erythema and slight edema were observed at the injection sites of all control group animals and all test group animals at which only Freund's adjuvant / 0 .9% aqueous NaCl-solution (1 : 1) was applied . Injection of 5% test substance preparations in olive oil DAB 10 or in Freund's adjuvant / 0 .9% aqueous NaCl-solution (1 : 1) caused well-defined erythema and slight edema in all test group animals. Olive oil DAB 10, which was applied to all control group 1 and 2 animals, caused well-defined erythema.

After the percutaneous induction with the undiluted test substance incrustation, partially open (caused by the intradermal induction) could be observed in addition to well-defined erythema and slight edema in all test group animals.

-Challenge:

The challenge with a 75% test substance preparation did not cause any skin reactions, neither in the control group 1 animals nor in the test group animals (24 and 48 hours after removal of the patch) .

Olive oil DAB 10, which was applied as a vehicle control to the test group and control group 1 and 2 animals, did not cause any skin reactions.

Since no borderline results were observed, a 2nd challenge was not performed.

-Body weights:

The expected body weight gain was generally observed in the course of the study.

Applicant's summary and conclusion

Interpretation of results:
other: Non-sensitizer
Conclusions:
The results of this study according to OECD guideline 406 show that Vinyl propionate (CAS 105-38-4) does not have a sensitizing effect on the skin of the guinea pig in the Maximization Test under the test conditions chosen.
Executive summary:

The substance Vinyl propionate (CAS 105-38-4) was tested for its sensitizing effect on the skin of the guinea pig in the Maximization Test based on the method of Magnusson and Kligman. The study was erformed from January 14, 1997 until February 07, 1997. The intradermal induction with 5% test substance preparations caused slight to well-defined signs of skin irritation in all test group animals.

After the percutaneous induction with the undiluted test substance incrustation, partially open (caused by the intradermal induction) could be observed in addition to well-defined erythema and slight edema in all test group animals. A challenge was performed 14 days after the percutaneous induction. The number of animals with skin findings after the challenge is summarized in the following table:

 

Challenge: test substance 75% in olive oil DAB 10   Challenge: olive oil DAB 10
 Control group 1 0/10   0/10
 Control group 2*  no application of test substance  0/10
 Test group  0/20  0/20

x/y: number of positive reactions/number of animals tested (reading at 24h and/or 48h after the removal of the patch)

*control group 2 that had been intended for a potential 2nd challenge was not treated with the test substance, since a 2nd challenge was not necessary on the basis of the unambiguous results of the lst challenge.

Based on the results of this study and applying the evaluation criteria it was concluded that the Vinyl propionate (CAS 105-38-4) does not have a sensitizing effect on the skin of the guinea pig in the Maximization Test under the test conditions chosen.