2-methylbutyraldehyde

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions (no individual caging, test substance concentrations were not in consecutive order)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

yes

Remarks:

: no individual caging, test substance concentrations were not in consecutive order

Test guideline 3

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

2-methylbutyraldehyde

EC Number:

202-485-6

EC Name:

2-methylbutyraldehyde

Cas Number:

96-17-3

Molecular formula:

C5H10O

IUPAC Name:

2-methylbutanal

Details on test material:

- Name of test material (as cited in study report): 2-methyl butyraldehyde
- Physical state: clear colorless liquid
- Analytical purity: 98.2 ± 0.06 %
- Impurities (identity and concentrations): 0.049 wt% water, 0.3%, 0.9% and 0.3% not identified substances A, B, and C
- Purity test date: no data
- Lot/batch No.: TK2632B-16OCT2007
- Stability under test conditions: stable, recertification date January 31, 2010
- Storage condition of test material: no data

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan, Indianapolis, IN, USA
- Age at study initiation: approx. 10 - 11 weeks
- Weight at study initiation:
- Housing: up to 6 per cage in filter tubs containing corncob bedding
- Diet (e.g. ad libitum): LabDiet Certified Rodent Diet #5002 (PMI Nutrition International, St. Louis, MO, USA) in pelleted form, ad libitum
- Water (e.g. ad libitum): municipal drinking water, ad libitum
- Acclimation period: at least 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 1
- Humidity (%): 40 - 70
- Air changes (per hr): 12 - 15 times/hour
- Photoperiod (hrs dark / hrs light): 12-hour light/dark

Study design: in vivo (LLNA)

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

5, 25, and 100 % test substance in vehicle

No. of animals per dose:

6

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: soluble in vehicle
- Application: one female mouse was used per concentration (1%, 5%, 25%, 50%, 75%, and 100% of test substance). Animals received one application of test solution on three consecutive days spread on the dorsal surface of each ear (25µL/ear) in a manner to prevent material loss
- Irritation: pure (100%) test substance caused slight erythema at day 3 which resolved by day 6
- Lymph node proliferation response: no data

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: local lymph node assay
- Criteria used to consider a response positive: a stimulation index (SI) of ≥ 3 (i.e., 3-fold greater proliferation than control animals) is considered positive for dermal sensitization potential of the test substance

TREATMENT PREPARATION AND ADMINISTRATION
- Test solutions:Test solutions were prepared daily just prior to dosing. Concentrations were not verified analytically.
- Administration: test material (25 µL/ear) was administered once daily for three consecutive days on the dorsal surface of both ears using an adjustable pipette as for the range finding test.

OBSERVATIONS
- Visual control of ears, evaluation of erythema: prior to application and on day 2, 3 and 6
- Weighing: on day 1 and 6

TREATMENT WITH 3H-THYMIDINE AND PREPARATION OF LYMPH NODE CELL SUSPENSION
- 3H-Thymidine treatment: on day 6 five hours prior to sacrifice, all mice received a 250 µL intravenous injection via the lateral tail vein containing 20 µCi of 3H-thymidine (specific activity 2Ci/mmol) diluted in phosphate buffered saline (PBS).
- Isolation of lymph nodes: both of the auricular lymph nodes per mouse were excised. Single cell suspensions in PBS were prepared by gentle mechanical disaggregation using a tissue homogenizer. Cells were washed twice and suspended in trichloroacetic acid for approx. 18 h. The resulting precipitate was separated by centrifugation and the radioactivity in each precipitate was measured using ß-scintillation counting.

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Statistics:

Means and standard deviation (SD) were generated for body weight data (absolute and gain) and the LLNA response (dpm & SI values).
Body weight and dpm data were analyzed by a one-way analysis of variance. When differences were indicated by the ANOVA, a comparison of treated vs. control groups was done using a Dunnett’s t-test. The alpha level at which all tests were conducted was 0.05.

Results and discussion

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: See table below. For the 5% and 25% group no significant response was observed. For the 100% group, the radioactivity count was increased about 4 fold. A SI of 4.4 was calculated. EC3 was calculated to be 70%.

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: See Table below

Any other information on results incl. tables

Results of the local lymph node assay (mean ± sd)

 

Concentration	DPM	SI
Solvent control	550.50 ± 351.43	1.0 ±0.6
5%	546.83 ± 399.41	1.0 ± 0.6
25%	494.17 ± 212.05	0.9 ± 0.4
100%	2466.4 ± 803.88*	4.4 ± 1.4
Positive control	4073.8 ± 1325.3*	7.4 ± 2.4

 

* statistically different; α = 0.05

 

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information

Conclusions:

In this LLNA, a positive response was only obtained after application of the highest dose (100% test substance). At this dose, local irritation reaction was observed. Test groups exposed to 5% and 25% test substance solution did not show any reaction. The 50% solution was not tested. Thus there is no information about an effect in the concentration range between 25% and 100% (additional value in dose-response-plot).

Executive summary:

In a local lymph node assay with 2-methylbutyraldehyde (2-methylbutanal) (purity 98.2%), adult female CBA/J mice (6 animals per group) were tested using concentrations of 5, 25, and 100% test substance in vehicle (4:1 acetone/olive oil). Hexyl cinnamic aldehyde (30% in vehicle) was used as positive control material.

 

The positive control substance displays the appropriate response. For 2-methylbutyraldehyde, there was no effect of treatment on body weight development. Skin irritation, which resolved in all mice by day 6, was only observed on day 3 in mice dosed with 100% test substance (slight to well-defined erythema, 4 and 2 of 6 mice respectively). Increases in lymph cells were only observed in the 100% test group. For this group a SI of 4.4 was determined. EC3 was 70%.

 

In this study, 2-methylbutanal is a weak dermal sensitizer (categorized according to the expert ECETOC panel - Technical Report No. 87, 2003) (Dow 2008).

 

This study is classified as acceptable. It was performed according to OECD test guideline 429 with some restrictions (no individual caging, test substance concentrations were not in consecutive order).