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Description of key information

Buehler OECD 406: Skin sensitizing 1B (BASF SE 2019)

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
September 2018 - January 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
yes
Type of study:
Buehler test
Justification for non-LLNA method:
Substanceclass is know to lead to false positive results in LLNA Test, therefore a Guineapig is conducted.
Specific details on test material used for the study:
For anayltical details see study report No. 18L00152
Species:
guinea pig
Strain:
Dunkin-Hartley
Remarks:
Dunkin Hartley, Crl:HA, SPF
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Wiga GmbH, Germany
- Females (if applicable) nulliparous and non-pregnant: yes
- Microbiological status of animals, when known: spf
- Age at study initiation: no specified
- Weight at study initiation: 250 g – 274 g
- Housing:

Type of cage:
Stainless steel wire mesh cages with grating with shallow cage body; floor area: 4225 cm2
Enrichment:
Wooden gnawing blocks (Type KNH E-041); Abedd ® Lab. and Vet. Service GmbH Vienna, Austria
Bedding:
H 15005-29; Ssniff, Spezialdiäten GmbH (Experimental Animal Diets Inc., 59494 Soest, Germany)
Number of animals per cage:
5

- Diet (e.g. ad libitum): FD1 SQC; SDS Special Diets Services, 67122 Altrip, Germany, ad libitum
- Water (e.g. ad libitum): Tap water ad libitum
- Acclimation period:Acclimatization period of at least 5 days before the beginning of the experimental phase


ENVIRONMENTAL CONDITIONS
- Temperature (°C):20 +- 3°C;
- Humidity (%):30 - 70%
- Air changes (per hr):10
- Photoperiod (hrs dark / hrs light):12h/12h
Route:
epicutaneous, occlusive
Vehicle:
polyethylene glycol
Remarks:
Polyethylene glycol 400
Concentration / amount:
1st induction (60 %): suspension (paste)
2nd and 3rd induction (50%): suspension (paste)
Day(s)/duration:
Exposure period: 6 hours
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
polyethylene glycol
Remarks:
PEG 400
Concentration / amount:
Concentration test item 50 % (w/w)
Day(s)/duration:
Duration of exposure: 6 hours
Readings: 24, 48 and 72 h after the removal of the patch
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
20
Details on study design:
RANGE FINDING TESTS:
In accordance with the cited guidelines a mild irritating concentration should be used in the main test for induction, whereas the maximum non-irritant concentration should be applied for challenge.
Amount applied: 0.5 g or 0.5 mL of the test item preparation was applied to each animal.
Occlusive dressing: 2 x 2 cm gauze patches (6 layers surgical gauze Ph. Eur. from Lohmann GmbH & Co. KG) containing the test item preparations were applied to the skin of the flanks under an occlusive dressing. The dressing consisted of rubberized linen patches (4 x 4 cm from Russka), patches of Idealbinde (4 x 4 cm from Pfälzische Verbandstoff-Fabrik) and Fixomull® Stretch (adhesive fleece) from Beiersdorf AG.
Duration of exposure: 6 hours
Application site: Right and left flank anterior and posterior
Application frequency: One application
Number of test animals: 3
Readings: 1, 24, 48 and 72 h after removal of the patch
The animals were distributed as follows:
Flank region
(application site) Concentration % Number of Animals per Group
Left anterior 60 3
Left posterior 50 3
Right anterior 25 3
Right posterior 10 3
In the pretest, the highest concentration that could be technically used was 60% (w/w).

MAIN STUDY
Number of animals of the control group:10
Number of animals of the test group:20
A. INDUCTION EXPOSURE
3 inductions were conducted (day 0, 7 and 14)
Amount applied: 0.5 g of the test item preparation were applied to each test group animal. The control group received 0.5 mL of the vehicle.
Occlusive dressing: 2 x 2 cm gauze patches (6 layers surgical gauze Ph. Eur. from Lohmann GmbH & Co. KG) containing the test item preparation or vehicle were applied to the skin of the flank under an occlusive dressing. The dressing consisted of rubberized linen patches (4 x 4 cm from Russka), patches of Idealbinde (4 x 4 cm from Pfälzische Verbandstoff-Fabrik) and Fixomull® Stretch (adhesive fleece) from Beiersdorf AG.
Duration of exposure: 6 hours
Application site: Intact flank
Test item preparation (test group): left flank anterior
Vehicle (control group): left flank anterior
Due to distinct skin irritation in the test group the patches were shifted to the middle part of the left flank during the third induction.
Concentration of test item: 1st induction: 60 % (w/w)
2nd and 3rd induction: 50 % (w/w)
Application frequency: Three times at weekly intervals; days 0, 7 and 14 on the same or shifted application area
Readings: 24 h after the removal of the patch
The control group was treated analogously to the test group but only with the vehicle without the test item.



B. CHALLENGE EXPOSURE
The challenge was carried out 13 days after the third induction.
Amount applied: 0.5 g of the test item preparation and 0.5 mL of the vehicle were applied to each animal.
Occlusive dressing: 2 x 2 cm gauze patches (6 layers surgical gauze Ph. Eur. from Lohmann GmbH & Co. KG) containing the test item preparation or vehicle were applied to the skin of the flank under an occlusive dressing. The dressing consisted of rubberized linen patches (4 x 4 cm from Russka), patches of Idealbinde (4 x 4 cm from Pfälzische Verbandstoff-Fabrik) and Fixomull® Stretch (adhesive fleece) from Beiersdorf AG.
Duration of exposure: 6 hours
Application site: Intact flank
Test item preparation: right flank posterior
Vehicle: left flank posterior
Concentration test item: 50 % (w/w)
Readings: 24, 48 and 72 h after the removal of the patch
The control group was treated analogously to the test group.

Positive control
A 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 controls with Alpha-Hexylcinnamaldehyde, techn. 85% showed that the test system was able to detect sensitizing compounds under the laboratory conditions chosen.
Positive control substance(s):
yes
Remarks:
A 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.
Reading:
other: Total
Hours after challenge:
48
Group:
test chemical
Dose level:
50%
No. with + reactions:
4
Total no. in group:
20
Remarks on result:
other: In total, 4/20 test group animals showed skin reactions after 24 and 48 hours after removal of the patches.
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
50%
No. with + reactions:
3
Total no. in group:
20
Clinical observations:
none
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
50%
No. with + reactions:
2
Total no. in group:
20
Clinical observations:
none
Interpretation of results:
Category 1B (indication of skin sensitising potential) based on GHS criteria
Executive summary:

"Confidential item" was tested for its sensitizing effect on the skin of the guinea pig in the BUEHLER Test based on the method of BUEHLER, E.V. (1965) and according to OECD Guideline 406 (1992), the council regulation (EC) No 440/2008 part B.6. (2008) and the U.S. EPA OPPTS 870.2600 (2003) and MAFF 8147, 2000 guideline.
The test item concentrations for the main test were selected based on the results of the pretest, in which 3 guinea pigs of the same animal strain in total were used. In the pretest, the highest concentration that could be technically used was 60% (w/w). Therefore, descending concentrations of 60, 50, 25 and 10% (w/w) in polyethylene glycol 400 were chosen.
Based on the data of the pretest, the first induction was performed with a 60% (w/w) test item preparation in polyethylene glycol 400. Due to distinct local skin findings after the first induction, the second and third induction was performed with a 50% (w/w) test item preparation in polyethylene glycol 400. Likewise, a 50% test item preparation in polyethylene glycol 400 was chosen for the challenge application.
The main study was performed using 1 control group, which consisted of 10 animals and 1 test group, which consisted of 20 animals.
The inductions were performed on days 0, 7 and 14.
A challenge was carried out 13 days after the last induction.
In the main study, the animals did not show any signs of systemic toxicity.
As expected, no local skin findings could be observed in the control group neither after the inductions nor the challenge.
In the test group, 13 animals (13/20) revealed discrete or moderate erythema (grade 1 or 2) after the first induction, while after the second induction these skin findings were observed in 18 animals (18/20) animals. 13 animals (13/20) showed discrete or moderate erythema after the third induction.
After the challenge 4 animals (4/20) in total showed the following skin findings:
 Discrete erythema (grade 1)


Due to the unambiguous findings of the challenge, a second challenge was not performed.
Based on the results of this study and applying the evaluation criteria cited in chapter 3.8, it was concluded that "confidential testitem" has a sensitizing effect on the skin of the guinea pig in the BUEHLER Test under the test conditions chosen.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)
Additional information:

There is one reliable study regarding skin sensitisation with C8-18AS TEA&Mg (CAS 85586-38-5) available.


Within this study "Confidential item" was tested for its sensitizing effect on the skin of the guinea pig in the BUEHLER Test based on the method of BUEHLER, E.V. (1965) and according to OECD Guideline 406 (1992), the council regulation (EC) No 440/2008 part B.6. (2008) and the U.S. EPA OPPTS 870.2600 (2003) and MAFF 8147, 2000 guideline.
The test item concentrations for the main test were selected based on the results of the pretest, in which 3 guinea pigs of the same animal strain in total were used. In the pretest, the highest concentration that could be technically used was 60% (w/w). Therefore, descending concentrations of 60, 50, 25 and 10% (w/w) in polyethylene glycol 400 were chosen.
Based on the data of the pretest, the first induction was performed with a 60% (w/w) test item preparation in polyethylene glycol 400. Due to distinct local skin findings after the first induction, the second and third induction was performed with a 50% (w/w) test item preparation in polyethylene glycol 400. Likewise, a 50% test item preparation in polyethylene glycol 400 was chosen for the challenge application.
The main study was performed using 1 control group, which consisted of 10 animals and 1 test group, which consisted of 20 animals.
The inductions were performed on days 0, 7 and 14.
A challenge was carried out 13 days after the last induction.
In the main study, the animals did not show any signs of systemic toxicity.
As expected, no local skin findings could be observed in the control group neither after the inductions nor the challenge.
In the test group, 13 animals (13/20) revealed discrete or moderate erythema (grade 1 or 2) after the first induction, while after the second induction these skin findings were observed in 18 animals (18/20) animals. 13 animals (13/20) showed discrete or moderate erythema after the third induction.
After the challenge 4 animals (4/20) in total showed the following skin findings:
 Discrete erythema (grade 1)


Due to the unambiguous findings of the challenge, a second challenge was not performed.
Based on the results of this study and applying the evaluation criteria, it was concluded that "confidential testitem" has a sensitizing effect on the skin of the guinea pig in the BUEHLER Test under the test conditions chosen.


It is concluded by the registrant that this effect is unusual and not typical for the group of alkylsulfates (see below for database on AS sensitization tests). The registrant trace back this behavior to the specific production process of the UVCB substance and maybe formed smaller imprurities. Normally AS are produced in water based solvent situation, this is not the case here which is assumed to may cause the unusual result here. Further the concentration used in the Buehler test was also untypical high due to the fact that the used solvent could be more easier evaporated and the substance shows also an variable corrosive behavior. Both factor may also act as confounders, as it is know for some sensitization test irritation leads to false positive results for some substance groups. Hence the positive finding in the sensitization is not regularly explainable but assumed to be based on the specific production process and the UVCB characteristic.


Because there is comprehensive database on skin sensitization of (mono constituent) alkyl sulfates available. One of the reliable studies proved alkyl sulfates to be sensitizing. Two positive LLNAs were disregarded as the reported effects were linked to the irritating properties of alkyl sulfates. The LLNA is known to tend to produce false positive results with some irritating chemicals “such as some surfactant type chemicals”, which is already stated in the OECD Guideline. In addition, sodium lauryl sulfate (C12AS Na) is recommended as an agent to induce local irritation in a method to assess skin sensitizing properties of chemicals (OECD guideline 406, Guinea Pig Maximisation Test). Therefore there is a lot of experience with sodium lauryl sulfate in sensitization studies and no evidence occurred that it shows sensitizing properties. Thus, skin sensitization by members of the alkyl sulfates is generally unlikely. The counter ion does not influence the skin sensitizing potential in a substantial way.


Anyhow investigation so far were not able to derive a explanation for this this positive skin sensitizing behaviour of the substance in a sensitisation test. Therefore the substance is classified as skin sensitizer 1B and disconnected concerning this behavior from the group of alkylsulfates also reasoned by the specific production process. 


 



Migrated from Short description of key information:
Skin sensitisation (Buehler - OECD 406): sensitising 1B

Justification for selection of skin sensitisation endpoint:
One reliable study available. Therefore chosen as key study.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

According to the classification criteria of Directive 67/548/EEC and Regulation (EC) No 1272/2008 the substance needs to be classified for skin sensitisation 1B. No data is available for respiratory sensitisation.

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