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Description of key information

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
1988
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Comparable to guideline study with acceptable restrictions. The study was conducted under GLP; although concentrations of induction were below irritation potential, the study is considered reliable and relevant, however not adequate for classification in isolation.
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
1981, page 57 ff.
Deviations:
yes
Remarks:
(no pretreatment with sodium lauryl sulphate)
Principles of method if other than guideline:
Test concentration used for induction was the highest non-irritating concentration and not the lowest concentration which causes mild-to-moderate skin irritation. The animals were not pretreated with sodium lauryl sulphate, as required by recent guidelines for testing of non-irritating substances. Furthermore, no positive control substance was tested in parallel and no periodically performed sensitivity and reliability check of the experimental technique is reported.
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Test material information:
Composition 1
Species:
guinea pig
Strain:
other: Pirbright / Hoe: DHPK(SPF-LAC.)/Boe
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Lippische Versuchtierzucht Hagemann GmbH & Co. KG , Hameiner Strasse 3, 4923 Exertal 1
- Age at study initiation: Not provided
- Weight at study initiation: 228-293 g
- Fasting period before study: Not provided
- Housing: max 5 guinea pigs/cage, Makrolon IV cages, (h 20 x b 30x l 55 cm)
- Diet (e.g. ad libitum): ad libitum, Ssniff -G® (Alleindiät für Meerschweinchen), Ssniff Spezialdiäten GmbH, 4770 Soest/Westfalen, pellets, 1.0 cm long, 0.5 cm diameter
- Water (e.g. ad libitum): ad libitum, Makrolon bottles (Firma Becker & Co., 4620 Castrop-Rauxel), tap water as for human use, analytical and bacteriological controls every half year
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 ± 2 °C (measured with thermohygrometer twice daily)
- Humidity (%): 50-85% (measured with thermohygrometer twice daily)
- Air changes (per hr): Not provided
- Photoperiod (hrs dark / hrs light): 12/12 (12hours daylight from 7.00-19.00h)

Route:
intradermal and epicutaneous
Vehicle:
other: deionised water
Concentration / amount:
No information on content of active ingredient of the test substance in this study report. However, according to producer information substance [Trade name] has 40 % act. ingr.
INDUCTION:
- intradermal treatment: 10 % test substance (corresponding to 4 % act.ingr.)
- epicutaneous treatment: 50 % test substance (corresponding to 20 % act.ingr.)
CHALLENGE: 50 % test substance (corresponding to 20 % act.ingr.)
Route:
epicutaneous, occlusive
Vehicle:
other: deionised water
Concentration / amount:
No information on content of active ingredient of the test substance in this study report. However, according to producer information substance [Trade name] has 40 % act. ingr.
INDUCTION:
- intradermal treatment: 10 % test substance (corresponding to 4 % act.ingr.)
- epicutaneous treatment: 50 % test substance (corresponding to 20 % act.ingr.)
CHALLENGE: 50 % test substance (corresponding to 20 % act.ingr.)
No. of animals per dose:
20 (10 male, 10 female)
Details on study design:
RANGE FINDING TESTS: In order to exclude primary skin irritation, two guinea pigs underwent single dermal treatment under occlusive conditions with the following concentrations (0.5 mL / animal): 100% (= undiluted), 75%, 50% and 10% solution in deionised water (corresponding to 40, 30, 20 and 4% act.ingr., respectively). An area of approx. 8x5 cm over the schoulders was clipped short. Two animals per concentration were treated.

MAIN STUDY
A. INDUCTION EXPOSURE first stage (intradermal)
- No. of exposures: 6 sites in 40 animals
- Exposure period: single injection
- Test group: 10 male and 10 female guinea pigs (1 group with 3 bilateral injections)
- Control group: 10 male and 10 female guinea pigs (1 group with 3 bilateral injections)
- Site: bilaterally to the spine starting in the cranio-dorsal area
- Frequency of applications: 1 (6 sites)
- Concentrations: - Test groups: 0.05 mL injection of 10% solution of the test substance in deionised water (2 injection sites bilaterally to the spine); 0.05 mL injection of 10% solution of the test substance in Freund Complete Adjuvans (2 injection sites bilaterally to the spine); 0.05 mL injection of Freund Complete Adjuvans undiluted (2 injection sites bilaterally to the spine)
- Control group: 0.05 mL injection of Freund Complete Adjuvans undiluted (2 injection sites bilaterally to the spine); 0.05 mL injection of 10% deionised water in Freund Complete Adjuvans (2 injection sites bilaterally to the spine); 0.05 mL injection of undiluted deionised water (2 injection sites bilaterally to the spine)

B.INDUCTION EXPOSURE second stage (epicutaneous)
- No. of exposures: 2 (x 6 sites) in 40 animals
- Day(s) of challenge: 7 days
- Exposure period: 48 hours occlusive dressing
- Test group: 20 animals
- Control group: 20 animals
- Site: the 3 sites bilaterally of the spine
- Concentrations: - Test group: 0.5mL of a 50% solution of the test substance in deionised water (corresponding to 20% act.ingr.)
- Control group: 0.5 mL of undiluted deionised water


C. CHALLENGE EXPOSURE 1
- No. of exposures: 2 (x 6 sites) in 40 animals
- Day(s) of challenge: 3 weeks (after intradermal injection)= 21 days
- Exposure period: 24 hours
- Test group: 20 animals
- Control group: 20 animals
- Site: the 3 sites bilaterally of the spine
- Concentrations: - Test group: 0.5mL of a 50% solution of the test substance in deionised water on the left side; 0.5 mL of undiluted deionised water on the right side
- Control group: 0.5mL of a 50% solution of the test substance in deionised water on the left side; 0.5 mL of undiluted deionised water on the right side

- Evaluation (hr after challenge): 24 h and 48h

Challenge controls:
yes: 0.5 mL of undiluted deionised water on the right side
Positive control substance(s):
no
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
50% test substance (corresponding to 20% act.ingr.)
No. with + reactions:
1
Total no. in group:
20
Clinical observations:
one animal showed a moderate erythema (reaction score 2)
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 50% test substance (corresponding to 20% act.ingr.). No with. + reactions: 1.0. Total no. in groups: 20.0. Clinical observations: one animal showed a moderate erythema (reaction score 2).
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
50 % test substance (corresonding to 20% act.ingr.)
No. with + reactions:
0
Total no. in group:
20
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 50 % test substance (corresonding to 20% act.ingr.). No with. + reactions: 0.0. Total no. in groups: 20.0.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
50 % test substance (corresponding to 20% act.ingr.)
No. with + reactions:
0
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 50 % test substance (corresponding to 20% act.ingr.). No with. + reactions: 0.0. Total no. in groups: 20.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
50% test substance (corresponding to 20% act.ingr.)
No. with + reactions:
1
Total no. in group:
20
Clinical observations:
one animal showed a slight erythema (reaction score 1)
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 50% test substance (corresponding to 20% act.ingr.). No with. + reactions: 1.0. Total no. in groups: 20.0. Clinical observations: one animal showed a slight erythema (reaction score 1).

Range finding: 100%: 24h and 48h : slightly erythema; 75%: 24h : very slightly erythema; 50% and 10%: no primary dermal irriatation

 Main test: Test group:24h after retesting (50% solution): 1 animal with moderate erythema. 48h after retesting all 20 animals of the test group had no reaction.                   

 Control group: after 24h no reaction on test site or on control site. At 48 h 1/20 animals showed a slight erythema (score 1) at the test substance treated skin area.

19 of the 20 test animals had no reaction, thus the test substance [Trade name] (50% solution) can be classified as “probably not sensitising”. The significant irritation in one test animal can still be assigned to chance.

Table 1. Range finding: individual observations

Animal number

Sex

Concentration

24h

48h

1

m

100% undiluted

1

0

2

f

100% undiluted

1

±

1

m

75% in deionised water

±

0

2

f

75% in deionised water

0

0

1

m

50% in deionised water

0

0

2

f

50% in deionised water

0

0

1

m

10% in deionised water

0

0

 

Table2. Individual observations after retesting (Attempt C) in the Test group (Right flank: 0.5 mL deionised water; Left flank: 0.5 mL test substance 50% solution in deionised water)

Animal number

Sex

24 hours

48 hours

Right flank

Left flank

Right flank

Left flank

1

M

0

0

0

0

2

M

0

0

0

0

3

M

0

0

0

0

4

M

0

0

0

0

5

M

0

2

0

0

6

M

0

0

0

0

7

M

0

0

0

0

8

M

0

0

0

0

9

M

0

0

0

0

10

M

0

0

0

0

11

F

0

0

0

0

12

F

0

0

0

0

13

F

0

0

0

0

14

F

0

0

0

0

15

F

0

0

0

0

16

F

0

0

0

0

17

F

0

0

0

0

18

F

0

0

0

0

19

F

0

0

0

0

20

F

0

0

0

0

 

Table3. Individual observations after retesting (Attempt C) in the Control group (Right flank: 0.5 mL deionised water; Left flank: 0.5 mL test substance 50% solution in deionised water)

Animal number

Sex

24 hours

48 hours

Right flank

Left flank

Right flank

Left flank

1

M

0

0

0

0

2

M

0

0

0

0

3

M

0

0

0

0

4

M

0

0

0

0

5

M

0

0

0

1

6

M

0

0

0

0

7

M

0

0

0

0

8

M

0

0

0

0

9

M

0

0

0

0

10

M

0

0

0

0

11

F

0

0

0

0

12

F

0

0

0

0

13

F

0

0

0

0

14

F

0

0

0

0

15

F

0

0

0

0

16

F

0

0

0

0

17

F

0

0

0

0

18

F

0

0

0

0

19

F

0

0

0

0

20

F

0

0

0

0

Interpretation of results:
other: probably not sensitizing
Remarks:
Criteria used for interpretation of results: expert judgment
Conclusions:
Based on current study, the test item can be classified as “probably not sensitising” as 19 of the 20 test animals had no reaction to the test substance (50% solution) according to the method of Magnusson and Kligman (OECD 406) and the distinct irritation in 1 test animal can still be assigned to chance.
Executive summary:

After an acclimation period of at least 7 days, 2 groups (test and control group) of 20 animals each were tested. Before treatment the animals were shaved in the shoulder area (approx. 8 x 5 cm). In order to exclude primary dermal irritation, 2 guinea pigs were tested in a single dermal (occlusive) application (0.5 mL (g) /animal) with following concentrations: 100% undiluted, 75%, 50% and 10% solution in deionised water of a test formulation containing 40% active ingredient (=range finding test).

In Attempt A (intradermal induction) each animal was injected in 2 injection sites (pairs) bilaterally to the spine (starting in the cranio-dorsal area) with 0.05 mL as follows:

Test group:1 pair (2) injections of test substance 10% solution in deionised water; 1 pair (2) injections of test substance 10% solution in FCA and 1 pair (2) injections of FCA undiluted.

Control group:1 pair (2) injections of FCA undiluted; 1 pair (2) injections of dionised water 10% solution in FCA and 1 pair (2) injections of deionised water undiluted.

In Attempt B , 7 days after intradermal induction, the first dermal induction started with 0.5 mL of the test substance (50% solution= maximal concentration with no primary irritation in range finding) on the left flank and 0.5 mL vehicle (deionised water) on the right flank in a closed patch test. After 48 hours the dressing was removed.

In Attempt C (Challenge), 3 weeks after intradermal treatment, the second single dermal application or challenge was executed with “Hill-Top” –Chambers, with 0.5 mL of the test substance (50% solution) on the left flank and 0.5 mL vehicle (deionised water) on the right flank. After 24 hours the dressing was removed. 24 Hours and 48 hours after removal evaluation was done.

The range finding showed no primary irritation in the 50% and 10% solution group.

 In Attempt C (Challenge) there was 1 animal in the test group (50% solution) with slightly erythema at 24h evaluation. At 48h after challenge none of the 20 animals of the test group had any reaction. In the control group after 24 and 48h there were no reactions on the test site or the control site. In 19 of the 20 test animals no reaction was observed, thus the test substance (50% solution or 20% active ingredient) can be classified as “probably not sensitising”. The significant irritation in 1 test animal can still be assigned to chance.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Skin sensitisation was assessed based on various Weight-of-Evidence studies from read-across substances:

- In a skin sensitisation study with read-across substance ' Butanedioic acid, sulfo-, 4-[2-[(2-hydroxyethyl)amino]ethyl] ester, N-C18-unsatd. acyl derivs., disodium salts' according to the method of Magnusson and Kligman, 2 groups (test and control group) of 20 animals each were tested with the registered substance containing 40% active ingredient (Sterner and Chibanguza, 1988).

The test substance (50% solution or 20% active ingredient) can be classified as “probably not sensitising”. The significant irritation in 1 test animal can still be assigned to chance.

-In a skin sensitisation study with read across substance 'Aspartic acid, N-(3-carboxy-1-oxo-sulfopropyl)-N-(C16-C18 (even numbered), C18unsaturated alkyl) tetrasodium salts', a 15 mm patch of the test item containing 35.8% active ingredient (2.5% in petrolatum) was applied to patch sites on the backs or volar forearms of 100 subjects for ten alternate-day 24 hour periods under occlusion (Kligman, 1977). Following a seven-day rest period, 15mm challenge patches of the test item (1% in petrolatum) were applied in the same manner to fresh sites on the backs or volar forearms of all 100 subjects for 24 hours. Challenge sites were read on removal of the patch and 24 hours thereafter, using the 0-4 scale. There were no instances of irritation or sensitization from this material on the Draize-Shelanski Patch Test. It is unlikely that this test item would present a danger of irritation or sensitization in normal, intended use.

Since the registered substance has a higher moleculr weght of the previous similar considered, it can be assumed that skin penetration is lower and skin sensitisation potential even reduced. it can be concluded that the registered substance is a non sensitizer


Migrated from Short description of key information:
Weight-of-evidence on sensitisation potential of registered substance and read across substances in a guinea pig maximisation model and human patch test all indicated there is no sensitisation potential.

Justification for selection of skin sensitisation endpoint:
Study performed on the analogous with the highest molecular weight, where skin adsorption can be more similar

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

European Directives concerning the classification, packaging and labelling of dangerous substances (Council Regulation (EC) No. 1272/2008 and subsequent revisions) the criteria for the classification of skin sensitizer are:

Category 1:

(a) if there is evidence in humans that the substance can lead to sensitisation by skin contact in a substantial number of persons; or

(b) if there are positive results from an appropriate animal test

Sub-category 1A: substances showing a high frequency of occurrence in humans and/or a high potency in animals can be presumed to have the potential to produce significant sensitisation in humans. Severity of reaction may also be considered.

Sub-category 1B: substances showing a low to moderate frequency of occurrence in humans and/or a low to moderate potency in animals can be presumed to have the potential to produce.

The results of the test performed on similar substances and skin adsorption considerations will assess the registered substance as not classified under the Regulation 1272/2008