Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline conform GLP study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Name of test substance: Hydroxypivalic acid neopentylglycol ester flakes
CAS No.: 1115-20-4

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Test system / strain / quality: Mouse / CBA/J
- Age on day 0: 8 – 12 weeks
- Sex: Female
- Supplier: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, 97633 Sulzfeld
- Acclimatization period: 14 days before the first test-substance application
- Body weight on day 0: 17.4 g – 21.5 g
- Housing: single in Makrolon cage, type II
- Diet (e.g. ad libitum): Kliba-Labordiät (Maus / Ratte Haltung “GLP”), Provimi Kliba SA, Kaiseraugst, Basel, Switzerland, ad libitum
- Water (e.g. ad libitum): Tap water; ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 – 24°C
- Humidity (%): 30 – 70%
- Photoperiod (hrs dark / hrs light): 12 h /12 h
- Bedding: Lignocel PS14; SSNIFF

Study design: in vivo (LLNA)

Vehicle:
propylene glycol
Concentration:
50 % ; The 50% preparation was the maximum technically applicable concentration. The study was carried out as a limit test, using 1 test group and 1 control group.
No. of animals per dose:
5
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: A solubility experiment was performed according to the recommendations given by OECD 429. The highest test-substance concentration, which can be technically used, was a 50% test-substance preparation.
- Irritation: none
- Lymph node proliferation response: none

MAIN STUDY
The study was carried out as a limit test, using 1 test group and 1 control group. Each test animal was applied with 25 μL per ear of the test-substance preparation to the dorsum of both ears for three consecutive days. The control group was treated with 25 μL per ear of the vehicle alone.
Three days after the last application the mice were injected intravenously with 20 μCi of 3H-thymidine in 250 μL of sterile saline into a tail vein. About 5 hours after the 3H-thymidine injection, the mice were sacrificed and the auricular lymph nodes were removed. The weights of each animal’s pooled lymph nodes were determined. Thereafter lymph nodes were pooled group wise and further evaluated by measuring their cellular content and 3H-thymidine incorporation into the lymph node cells (indicators of cell proliferation). Moreover, a defined area with a diameter of 0.8 cm was punched out of the apical part of each ear and for each test group the weight of the pooled punches was determined in order to obtain an indication of possible skin irritation.
The stimulation indices (fold of change as compared to the vehicle control) for cell count, 3H-thymidine incorporation, lymph node weight and ear weight were measured. The increase SI of cell count by a factor of ≥ 1.5 and/or of 3H-thymidine incorporation by a factor of ≥ 3 as compared to the concurrent vehicle control group is generally considered as indicating a sensitizing potential of a test substance.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
mean velues, and standard diviation

Results and discussion

Positive control results:
A concurrent positive control (reliability check) with a known sensitizer was not included into this study. Studies using the positive control substance Alpha-Hexylcinnamaldehyde, techn. 85% are performed twice a year in the laboratory in order to show that the test system is able to detect sensitizing compounds under the test conditions chosen. Positive controls in these experiments gave the expected results.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: 0.83 (compared to the concurrent control) 2 .04 (compared to the historical control)
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: vehicle propylene glycol 1,380.9 (SI =1); historical control (mean value) 561.2 50% test substance in propylne glycol: 1,143.3

Any other information on results incl. tables

CELL COUNT, 3H-THYMIDINE INCORPORATION AND LYMPH NODE WEIGHT:

TEST GROUP MEAN VALUES AND STIMULATION INDICES

 

Test group

Treatment

Cell Counts

 

 

[Counts/Lymph Node Pair]

Stimulation Index

1

vehicle propylene glycol

historical control (mean)

8 634 667

6 809 867

1.00

2

50% in propylene glycol

7 776 000

0.90

1.14

 

Test group

Treatment

³H-thymidine incorporation

 

 

[DPM/Lymph Node Pair]

Stimulation Index

1

vehicle propylene glycol

historical control (mean)

1 380.9

561.2

1.00

2

50% in propylene glycol

1 143.3

0.83

2.04

 

Test group

Treatment

Lymph Node Weight

 

 

 

[mg/Lymph Node Pair]

Stimulation Index

1

vehicle propylene glycol

5.4

1.00

2

50% in propylene glycol

4.8

0.90

 

EAR WEIGHT: TEST GROUP MEAN VALUES AND STIMULATION INDICES

Test group

Treatment

Ear weight

 

 

[mg/animal]

Stimulation Index

1

vehicle propylene glycol

32.3

1.00

2

50% in propylene glycol

34.5

1.06

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information