Registration Dossier

Administrative data

Description of key information

Skin sensitisation: LLNA, mouse (CBA/Ca (CBA/CaOlaHsd)), female, IPGE, OECD 429, EU method B.42, GLP: not sensitising

Skin sensitisation: Modified Maguire method, Guinea pig, IPGE: not sensitising

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:
other: Well-documented OECD 429 guideline study under GLP without deviations on the registered substance itself.
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
OECD Guideline for the Testing of Chemicals No. 429 "Skin Sensitization: Local Lymph Node Assay" (adopted 22 July 2010)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
Method B42 Skin Sensitization (Local Lymph Node Assay) of Commission Regulation (EC) No. 440/2008
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
The Department of Health of the Government of the United Kingdom
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
other: CBA/Ca (CBA/CaOlaHsd)
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS B.V., Inc., Horst, The Netherlands
- Age at study initiation: 8 to 12 weeks
- Weight at study initiation: 15 to 23 g
- Housing: The animals were housed in suspended solid floor polypropylene cages furnished with softwood woodflakes. The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.
- Diet (e.g. ad libitum): 2014C Teklad Global Rodent diet supplied by Envigo RMS (UK) Limited, Oxon, UK, ad libitum
- Water (e.g. ad libitum): mains tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25 °C
- Humidity (%): 30 to 70%
- Air changes (per hr): approx. 15
- Photoperiod (hrs dark / hrs light): hours continuous light and 12 hours darkness
Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
undiluted or 50% or 25% v/v
No. of animals per dose:
4
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: For the purpose of the study, the test item was used undiluted and freshly prepared as a solution in acetone/olive oil 4:1. This vehicle was chosen as it produced the most suitable formulation at the required concentration.
The test item was formulated within 2 hours of being applied to the test system. It is assumed that the formulation was stable for this duration.
No analysis was conducted to determine the homogeneity, concentration or stability of the test item formulation. This is an exception with regard to GLP and has been reflected in the GLP compliance statement.
- Irritation: As no toxicological information was available regarding the systemic toxicity/irritancy potential of the test item, a preliminary screening test was performed using one mouse. The mouse was treated by daily application of 25 μL of the undiluted test item to the dorsal surface of each ear for three consecutive days (Days 1, 2, 3). The mouse was observed twice daily on Days 1, 2 and 3 and once daily on Days 4, 5 and 6. Local skin irritation was scored daily. Any clinical signs of toxicity, if present, were also recorded. The body weight was recorded on Day 1 (prior to dosing) and on Day 6.
- Lymph node proliferation response: The thickness of each ear was measured using a Mitutoyo 547-300S gauge (Mitutoyo Corporation), pre-dose on Day 1, post dose on Day 3 and on Day 6. Any changes in the ear thickness were noted. Mean ear thickness changes were calculated between time periods Days 1 and 3 and Days 1 and 6. A mean ear thickness increase of equal to or greater than 25% was considered to indicate excessive irritation and limited biological relevance to the endpoint of sensitization.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response: The test item will be regarded as a sensitizer if at least one concentration of the test item results in a threefold or greater increase in 3HTdR incorporation compared to control values. Any test item failing to produce a threefold or greater increase in 3HTdR incorporation will be classified as a "non-sensitizer".

TREATMENT PREPARATION AND ADMINISTRATION:
Test Item Administration
Groups of four mice were treated with the undiluted test item or the test item at concentrations of 50% or 25% v/v in acetone/olive oil 4:1. The preliminary screening test suggested that the test item would not produce systemic toxicity or excessive local skin irritation at the highest suitable concentration. The mice were treated by daily application of 25 μL of the appropriate concentration of the test item to the dorsal surface of each ear for three consecutive days (Days 1, 2, 3). The test item formulation was administered using an automatic micropipette and spread over the dorsal surface of the ear using the tip of the pipette.
A further group of four mice received the vehicle alone in the same manner.

3H-Methyl Thymidine Administration
Five days following the first topical application of the test item or vehicle (Day 6) all mice were injected via the tail vein with 250 μL of phosphate buffered saline (PBS) containing 3H-methyl thymidine (3HTdR: 80 μCi/mL, specific activity 2.0 Ci/mmoL, ARC UK Ltd) giving a total of 20 μCi to each mouse.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Positive control results:
A group of five animals was treated with 50 μL (25 μL per ear) of α-Hexylcinnamaldehyde, tech., 85% as a solution in acetone/olive oil 4:1 at a concentration of 25% v/v. The stimulation index was tested to be 6.08. α-Hexylcinnamaldehyde, tech., 85% was considered to be a sensitizer under the conditions of the test.
Parameter:
SI
Remarks on result:
other: Vehicle: n/a 25% test item: 0.72 50% test item: 1.07 undiluted test item: 2.44
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: Vehicle: dpm: 4324.08 / dmp/node: 540.51 25% test item: dpm: 3097.45 / dmp/node: 387.18 50% test item: dpm: 4625.22 / dmp/node: 578.15 undiluted test item: dpm: 10566.20 / dmp/node: 1320.78
Interpretation of results:
GHS criteria not met
Conclusions:
The study was conducted under GLP according to OECD guideline 429 on the registered substance itself. The method is to be considered scientifically reasonable with no deficiencies in documentation or any deviations, the validity criteria are fulfilled, positive and negative controls gave the appropriate response. Hence, the results can be considered as reliable to assess the sensitizing potential of 2,3-epoxypropyl isopropyl ether in mice. No clinical signs of toxicity were noted at a concentration of 100%, the SI determined after application of the undiluted test item was 2.44, which is below the threefold or greater increase in 3HTdR incorporation triggering classification. Hence, no classification as skin sensitizer is triggered.
Executive summary:

A dermal sensitization study was performed to assess the skin sensitization potential of 2,3-epoxypropyl isopropyl ether in the CBA/Ca strain mouse following topical application to the dorsal surface of the ear.

 

Following a preliminary screening test in which no clinical signs of toxicity were noted at a concentration of 100%, this concentration was selected as the highest dose investigated in the main test of the Local Lymph Node Assay. Three groups, each of four animals, were treated with 50 µL (25 µL per ear) of the undiluted test item or the test item as a solution in acetone/olive oil 4:1 at concentrations of 50% or 25% v/v. A further group of four animals was treated with acetone/olive oil 4:1 alone.

 

The Stimulation Index expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control group are as follows:

25%: SI = 0.72, negative

50%: SI = 1.07, negative

100%: SI = 2.44, negative

 

2,3-Epoxypropyl isopropyl ether was considered to be a non-sensitizer under the conditions of the test.

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

There is an in vivo Klimisch 1 GLP OECD 429 guideline study available, which assess the skin sensitization potential of 2,3-epoxypropyl isopropyl ether in the CBA/Ca strain mouse following topical application to the dorsal surface of the ear. Following a preliminary screening test in which no clinical signs of toxicity were noted at a concentration of 100%, this concentration was selected as the highest dose investigated in the main test of the Local Lymph Node Assay. Three groups, each of four animals, were treated with 50 µL (25 µL per ear) of the undiluted test item or the test item as a solution in acetone/olive oil 4:1 at concentrations of 50% or 25% v/v. A further group of four animals was treated with acetone/olive oil 4:1 alone.  The Stimulation Index expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control group are as follows:

25%: SI = 0.72, negative

50%: SI = 1.07, negative

100%: SI = 2.44, negative

2,3-Epoxypropyl isopropyl ether was considered to be a non-sensitizer under the conditions of the test. There is no indication given that the result is not relevant for human risk assessment, even though no human data is available, as all tested concentrations gave a clear negative result. The tonnage-driven data requirements under REACH are fully met, no data gaps were identified.

The supporting study confirms the negative result: A publication is available and regarding to this, for the test substance no skin sensitising effect could be detected in a modified Maguire test in the Guinea pig. (Reference : Rao, K. S., J. E. Betso, K. J. Olson: "A collection of guinea pig sensitization test results grouped by chemical class", Drug Chem. Toxicol. 4, 331 (1981))

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

In the available valid GLP OECD 429 guideline study, the test substance was identified as a non-sensitizing agent in the Local Lymph Node Assay, so no classification as skin sensitizer is triggered.