Diisopentyl ether

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was performed according to the OECD guideline No.429 (July 2010) and in compliance with the GLP guidelines.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

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)

Test material

Test material information

Constituent 1

Reference substance name:

Diisopentyl ether

EC Number:

208-857-4

EC Name:

Diisopentyl ether

Cas Number:

544-01-4

Molecular formula:

C10H22O

IUPAC Name:

3-methyl-1-(3-methylbutoxy)butane

Details on test material:

- Name of test material (as cited in study report): Diisopentyl ether
- Analytical purity: 99.2%
- Lot/batch No.: TAP 625

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source : Harlan Laboratories B.V., The Netherlands
- Age at study initiation: 8-9 weeks old
- Weight at study initiation: 19.8 g (+/- 1.1 g)
- Housing: group in Makrolon Type II/III cages with wire mesh top containing granulated soft wood bedding.
- Diet (e.g. ad libitum): free access to pelleted standard diet (Harlan Laboratories B.V., The Netherlands).
- Water (e.g. ad libitum): free access to tap water.
- Acclimation period: at least 5 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C (+/- 2°C)
- Humidity (%): 30-65%
- Photoperiod (hrs dark / hrs light): 12h/12h

Study design: in vivo (non-LLNA)

Induction

Concentration / amount:

Not applicable

Challenge

Concentration / amount:

Not applicable

No. of animals per dose:

Not applicable

Details on study design:

Not applicable

Challenge controls:

Not applicable

Study design: in vivo (LLNA)

Vehicle:

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

Remarks:

(Acetone, batch No. K40835814 (Merck, Darmstadt, Germany); Olive oil, batch No. 0001428703 (Sigma/Aldrich Chemicals, Taufkirchen, Germany))

Concentration:

For the preliminary test the concentrations were 50 (w/v) and 100% of the test item.
For the main test the concentrations were 0, 25, 50 (w/v) and 100% of the test item.

No. of animals per dose:

For the preliminary test: 1 female/dose (no control): left ear and right ear were treated with 50 and 100% of the test item.
For the main test: 4 females/dose and 4 females for the negative control (total of 16 animals)

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: The highest test item concentration, which could be technicallly used was 100% of the undiluted test item. The test item was soluble in the first recommended vehicle, acetone/olive oil (4/1, v/v). A solution was obtained at the maximum tested concentration of 50%.
- Irritation: Measurement of the ear thickness (using a micrometer) was performed prior to the first application (day 1), on day 3 and before sacrifice (day 6). Clinical signs and signs of local skin irritation were recorded daily. On day 4 to 6, the animals treated with 100% showed an ear erythema (score 1), in the animal treated with 50% this was only observed on day 4 of the pre-test. No signs of systemic toxicity were observed. Based on these results, the test item in the main study was assessed at 25, 50 (w/v) and 100%.
- Lymph node proliferation response: no measurement

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Lymph node cell proliferative responses were measured as described by Kimber and Dearman (1991).
- Criteria used to consider a positive response: The results were expressed as disintegration per minute (dpm) per group and per node. Stimulation indices (SI) were calculated according to the following formula: SI = dpm of treated group / dpm of control group. The test item was considered as a skin sensitizer when the SI for a dose group is higher than or equal to 3 and the data are compatible with a conventional dose response.
TREATMENT PREPARATION AND ADMINISTRATION:
The test item was prepared in the vehicle at the chosen concentrations. All dosage form preparations were made freshly on the day of the administration and any unused material was discarded that same day. On days 1, 2 and 3, a dose-volume of 25 μL of the control or dosage form preparations was applied to the dorsal surface of both ears.

Positive control substance(s):

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

Statistics:

The EC3 value was calculated according to the equation:
EC3 = (a-c)[(3-d)/(b-d)]+c
where EC3 is the estimated concentration of the test item required to produce a 3-fold increase in draining lymph node cell proliferative activity; (a,b) and (c,d) are respectively the co-ordinates of the two pair of data lying immediately above and below the S.I. value of 3 on the local lymph node assay dose response plot.

Results and discussion

Positive control results:

In the positive control group given HCA at the concentration of 25%, a stimulation index exceeding the threshold value of 3 (SI=9.03) was noted.
The study was therefore considered valid.

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

In this study Stimulation Indices (S.I.) of 1.74, 5.39 and 9.68 were determined with the test item at concentrations of 25 and 50% (w/v) in acetone:olive oil (4 +1 v/v) and 100% (undiluted test item), respectively. A clear dose response was observed. The EC3 value calculated was 33.6%.

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: See Table below (7.4.1).

Any other information on results incl. tables

All treated animals survived the scheduled study period and no signs of systemic toxicity or local skin irritation were observed. The results of the main test are presented in the Table below.

Table 7.4.1: Results of the main test

Test item concentration % (w/v)	Group	DPM (after correction of background)	Number of lymph nodes	DPM per lymph node	S.I.
0	1	1754	8	219.2	1.00
25%	2	3055	8	381.8	1.74
50%	3	9450	8	1181.2	5.39
100%	4	16981	8	2122.6	9.68

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The test item Diisopentyl ether was found to be a skin sensitizer under the test conditions of the study.

Executive summary:

In a local lymph node assay (LLNA) performed according to the OECD guideline No.429 and in compliance with the GLP guidelines, diisopentylether (purity 99.2%) dissolved in acetone/olive oil was assessed for its possible contact allergenic potential. The test item was administered to mice at concentrations of 25, 50% (w/v) and 100%.

The animals did not show any signs of systemic toxicity or local skin irritation during the course of the study and no cases of mortality were observed.

Stimulation Indices (S.I.) of 1.74, 5.39 and 9.68 were determined with the test item at concentrations of 25 and 50% (w/v) in acetone:olive oil (4 +1 v/v) and 100% (undiluted test item), respectively. A clear dose response was observed. The EC3 value calculated was 33.6%.