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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
3-10 September 2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Cross-reference
Reason / purpose:
read-across: supporting information
Reference
Endpoint:
skin sensitisation, other
Remarks:
Read-across
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: The information is derived from read across
Justification for type of information:
The full read-across document can be found in the Endpoint Summary and also the accompanying files.

Reason / purpose:
read-across source
Key result
Parameter:
EC3
Remarks:
%
Value:
25
Interpretation of results:
other: Skin sensitising (category 1B)
Remarks:
According to Regulation (EC) No. 1272/2008 and its amendments.
Conclusions:
Based on the data available it can be concluded that the substance is a skin sensitiser.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

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

In vivo test system

Test animals

Species:
mouse
Strain:
other: CBA/J
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Jackson Laboratories, Bar Harbor, ME 04609
- Age at study initiation: 9 weeks
- Specification: purpose-bred and experimentally naive at the outset of the study
- Weight at study initiation: 18 - 24 g (on initial dose day)
- Housing: Group housed 5 per cage
- Diet: Free access to Harlan Teklad Certified Rodent Chow 7012C
- Water: Free access to tap water
- Acclimation period: 7 days.

ENVIRONMENTAL CONDITIONS (target ranges)
- Temperature (°C): 22 - 25.6
- Humidity (%): 21 - 48
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:
other: EtOH/DEP
Concentration:
The test item at concentrations of 1, 2.5, 5, 10 and 25% w/v in the vehicle.
No. of animals per dose:
5
Details on study design:
A range finding test was not conducted for this study. The doses were selected so that the highest concentration maximizes exposure while avoiding systemic toxicity and excessive local irritation. Doses were selected based on known reported uses of the material.

TREATMENT PROCEDURES:
TOPICAL APPLICATION:
On Days 1, 2 and 3, each test animal in its group received an open application of 25 ul of an appropriate dilution of test item in vehicle to the dorsum of both ears. The positive control group (5 females) was treated with hexylicinnamaldehyde. All test and control animals were given a two-day rest period on Days 4 and 5.

ADMINISTRATION OF 3H-METHYL THYMIDINE:
On Day 6 of the study, all test and control animals were injected i.v. with 20 uCi of 3H-Thymidine in sterile saline. Five hours after injection, animals were sacrificed and the draining auricular lymph nodes excised.

DETERMINATION OF INCORPORATED 3HTdR:
The lymph nodes from each group were pooled and a single cell suspension was prepared. Cells were washed twice with PBS and precipitated with 5% trichloroacetic acid overnight at 2-8°C. The pellets were resuspended in 1 mL of TCA and transferred to a vial containing scintillation fluid. Incorporation of tritiated thymidine was measured by liquid scintillation counter.

OBSERVATIONS:
Individual body weights were recorded on Day 1 prior to dosing and Day 6 prior to injection. All test and control animals were observed daily for mortality and any excessive irritation at the test site. Clinical observations ere performed daily on day 4-6.

Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
DPM for each group was determined. Increases in 3H-thymidine incorporation relative to the vehicle-treated control was derived for each group.

Results and discussion

Positive control results:
The positive control item, hexyl cinnamic aldehyde, at 5%, 15% and 35% gave a Stimulation Index of 5.5, 6.1 and 19.5, respectively.

In vivo (LLNA)

Resultsopen allclose all
Key result
Parameter:
EC3
Remarks:
%
Value:
25
Parameter:
SI
Remarks on result:
other: The SI values calculated for the substance concentrations 1, 2.5, 5, 10 and 25% were 1.1, 1.9, 2.2, 2.1 and 3.0, respectively.

Any other information on results incl. tables

Results:

Animal group

Test item concentration

Average DPM count

SI (test / control ratio)

Results

Vehicle control

-

965.8

 

-

Test group I

1%

1069.7

1.1

-

Test group II

2.5%

1843.3

1.9

-

Test group III

5%

2160.2

2.2

-

Test group IV

10%

2001.4

2.1

-

Test group V

25%

2910.7

3.0

+

Positive control I

5%

5273.2

5.5

+

Positive control II

15%

5936.3

6.1

+

Positive control III

35%

18853.7

19.5

+

VIABILITY / MORTALITY:

There was no mortality throughout the study.

CLINICAL SIGNS:

All animals appeared normal for the duration of the study. No erythema or edema was seen in any of the animals during the study. On day 3, three out of 5 mice from the vehicle control group and 2 out of 5 mice from the 25% test material treatment group had ears that appeared wet. Furthermore, on days 2 -5, mice in the positive control groups had ears that appeared wet.

BODY WEIGHTS:

There were no statistically significant differences observed between any of the treatment groups.

Applicant's summary and conclusion

Interpretation of results:
other: Skin sensitising (category 1B)
Remarks:
According to Regulation (EC) No. 1272/2008 and its amendments.
Conclusions:
The SI values calculated for the substance concentrations 1, 2.5, 5, 10 and 25% were 1.1, 1.9, 2.2, 2.1 and 3.0, respectively. These results show that the test substance could elicit a SI ≥ 3. An EC3 has been derived resulted in an EC3 of 25%. The test substance was considered to be a sensitiser under the conditions of the test.
Executive summary:

The skin sensitisation potential of the substance has been tested according to OECD TG 429 and GLP principles. At 1, 2.5, 5, 10 and 25% the substance showed SI values of 1.1, 1.9, 2.2, 2.1 and 3.0, respectively. Reliable negative and positive controls were included. All animals appeared normal for the duration of the study.

These results show that the test substance could elicit a SI ≥ 3. An EC3 has been derived of 25%.

Based on the results, the substance is considered to be a skin sensitiser.