Registration Dossier

Administrative data

Description of key information

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
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:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
other: CBA/Ca
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River UK Limited, Kent, England
- Age at study initiation: 7-8 weeks
- Weight at study initiation: 16-19 g
- Housing: in pairs in suspended cages (dimensions 36.5 x 20.7 x 14 cm) with stainless steel grid tops and solid bottoms, bedding: wood shavings and nesting material (Nestlets, supplied by Datesand Limited Manchester, UK)
- Diet: Rat and Mouse No. 1 Maintenance Diet ad libitum, supplied by Special Diets Services Limited, Essex, UK
- Water: Water from the domestic mains supply ad libitum
- Acclimation period: 8 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-22
- Humidity (%): 55 (mean)
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2003-06-25 To: 2003-07-02
Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
10, 25 and 50%
No. of animals per dose:
4
Details on study design:
RANGE FINDING TESTS:
not performed, dose levels were selected based on existing data from a test on the test substance, produced under an old production process

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: (3H)-methyl thymidine
- Criteria used to consider a positive response: stimulation index (SI) >=3, together with consideration of dose-response and, where appropriate, statistical significance

TREATMENT PREPARATION AND ADMINISTRATION:
For 3 consecutive days, animals received open application of 25 µl of the appropriate formulation onto the dorsum of each ear. There were no treatment on Days 4 and 5. On Day 6 each animal received an intravenous injection of 250 µl of phosphate buffered saline (PBS) containing 20 µCi of 3H-methyl thymidine into the lateral tail vein. Approx. 5 h after intravenous administration animals were killed by exposure to carbon dioxide and exsanguinated. Each pair of draining auricular lymph nodes was collected from each animal. A single cell suspension of lymph node cells from each group was prepared by gentle mechanical desegregation through 200 µm mesh stainless steel gauze. The lymph node cells were washed twice in an excess of PBS and precipitated with 5% trichloroacetic acid at approx. 4°C for 18 h. The pellets were resuspended and solubilised in 200 µl soluene. Scintillation fluid (10 ml) was added and incorporation of tritiated thymidine measured by ß-scintillation counting as disintegrations per minute (dpm).
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
no statitistics performed
Positive control results:
The methods used on this study were periodically checked in the testing facility. Stimulation indices in recent positive control studies were 2.3, 5.5 and 7.5 in March 2003 and 1.9, 4.3 and 11.7 in May 2003 at concentrations of 10, 20 and 40% hexyl cinnamic aldehyde, thereby indicating that the test method employed is valid.
Key result
Parameter:
SI
Value:
1
Test group / Remarks:
10 % (new batch)
Key result
Parameter:
SI
Value:
2.6
Test group / Remarks:
25 % (new batch)
Key result
Parameter:
SI
Value:
1.7
Test group / Remarks:
50 % (new batch)
Key result
Parameter:
SI
Value:
1.9
Test group / Remarks:
50 % (old batch)

No adverse clinical signs were noted at any dose level in this study.

Body weight gain for animals receiving test substance was similar to controls.

Interpretation of results:
GHS criteria not met
Conclusions:
The target substance and Category Member 2 was considered not to be a sensitiser in mice.
Executive summary:

Delayed contact hypersensitivity potential of the target substance was investigated using CBA/Ca mice. The study was performed according to OECD guideline 429 and OECD Principles of Good Laboratory Practice.

Three groups of 4 female mice received an open application of 25µl of test substance onto the dorsum of each ear at concentrations of 10%, 25% or 50%. Another group of 4 females received only the vehicle, acetone:olive oil (4:1, v/v). Treatment was for 3 consecutive days. Three days later each animal received an intravenous injection of 3H-methyl thymidine, and 5 h later the draining lymph nodes were collected and the incorporation of tritiated thymidine assessed by scintillation counting.

On comparing the groups treated with the test substance with the control group, the Stimulation Indices were calculated to be 1.0, 2.6 and 1.7 for 10%, 25% and 50%, respectively.

Under the conditions of the study, the test substance was considered not to be a sensitiser in mice.

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

A mouse LLNA assay has been conducted at Huntingdon Life Sciences (HLS) with 2 -butyloctanoic acid at test substance concentrations of 25%, 50%, and 100%, respectively, using the “pooled treatment group approach” [HLS, Report No.: CND 007/032987/LN, 2003]. Significant toxicity including mortality was observed at the highest concentration. Remaining animals of the group were sacrificed before dosing at 3 day of the study for animal welfare reasons. In the low and mid dose groups greasy fur was observed post-dose from day 1. These signs had disappeared in the control and the low dose group, but were still present in animals of the mid dose group.  

The stimulation indices for the 25% and 50% dose groups were found to be 1.6 and 5.8, respectively. Since marked toxicity including mortality was observed in the highest dose groups and still some signs of toxicity in the mid dose groups the results were regarded to be inconclusive (Klimisch 4). To further address the sensitisation potential of the test item an additional LLNA was initiated using 2 different batches of the material.

The additional mouse LLNA was performed at Inveresk Research using 2 different batches of 2.butyloctanoic acid. For the “old” batch, that was already tested at HLS one single concentration (50%) was used, whereas for the “new” batch test substance concentrations of 10, 25, and 50%, respectively, were used. No signs of toxicity were observed in this study.

The stimulation indices were found to be 1.88 (50%, old batch) and 1.03, 2.60, and 1.72 for the 10%, 25%, and 50% dose groups of the “new” batch, respectively. Under the conditions of the test 2 -butyloctanoic acid was found to be “not sensitising”.

Conclusion: Positive responses have only been observed in one test at a concentration of 50%. At this concentration clinical signs (greasy fur) were present. At a higher concentration significant toxicity, including mortality was observed. In another test 2 different batches of the test item showed no signs of sensitisation up to the highest concentration of 50%. Taking into account the lack of structural alerts for sensitisation and lack of effects in one study using 2 different batches of the test item lesser weight should be laid to the single positive result in one test using a concentration of 50%. 2 -butyloctanoic acid therefore can be considered to be not sensitising.


Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

2 -Butyloctanoic acid based on the available information on sensitisation is not sensitising to the skin and does not require classification according Regulation (EC) No 1272/2008.