2-methoxyethyl acrylate

Endpoint summary

Description of key information

Skin sensitisation (OECD 429): 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

Study period:

25 May - 26 June 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP - Guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

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: Harlan Laboratories UK Ltd., Oxon, UK
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 15-23 g
- Housing: the animals were individually housed in suspended solid-floor polypropylene cages furnished with softwood wood flakes.
- Diet: 2014C Teklad Global Rodent diet (Harlan Laboratories UK Ltd., Oxon, UK)
- Water: tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-25
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

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

Concentration:

25 and 50% (v/v), 100% (undiluted)

No. of animals per dose:

preliminary screening test: 1
main assay: 4

Details on study design:

RANGE FINDING TESTS: in a preliminary screening test, the systemic toxicity/irritancy potential of the test item was assessed in each one mouse treated once daily with 25 µL of the undiluted test item (100%) and the test item at concentrations of 50%, 25% or 10% v/v in acetone/olive oil 4:1 on the dorsal surface of each ear for three consecutive days (Days 1, 2, 3). The mice were observed twice daily on Days 1, 2 and 3 and once daily on Days 4, 5 and 6. Local skin irritation was scored daily and any clinical signs of toxicity, if present, were also recorded. The bodyweight of each mouse was recorded on Day 1 (prior to dosing) and on Day 6. The thickness of each ear was measured using an Oditest micrometer (Dyer, PA), pre-dose on Day 1, post dose on Day 3 and on Day 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 sensitisation.

- Compound solubility: the test substance at 50% (v/v) in acetone/olive oil 4:1 was well soluble, and thus suitable for dosing.
- Irritation: no irritation indicated by an equal to or greater than 25% increase in mean ear thickness was noted. Very slight erythema was noted in animals treated with the test item at concentrations of 25% and 10% v/v in acetone/olive oil 4:1 during Days 3-5. No visual local skin irritation was noted in animals treated with the undiluted test item or the test item at a concentration of 50% v/v in acetone/olive oil 4:1.
- Clinical signs: no signs of systemic toxicity were observed.
Based on this information, the undiluted test item and the test item at concentrations of 50% and 25% v/v in acetone/olive oil 4:1 were selected for the main test.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: ³H-methyl thymidine (³HTdR) incorporation determined by β-scintillation
- Criteria used to consider a positive response: the test item will be regarded as a sensitiser if at least one concentration of the test item results in a 3-fold or greater increase in ³HTdR incorporation compared to control values. Any test item failing to produce a 3-fold or greater increase in ³HTdR incorporation will be classified as a "non-sensitiser".

TREATMENT PREPARATION AND ADMINISTRATION: based on the preliminary screening test, groups of 4 mice were treated with the undiluted test item (100%) or the test item at concentrations of 50% or 25% v/v in acetone/olive oil 4:1. The test substance (25 µL) was daily applied to the dorsal surface of each ear for three consecutive days (Days 1, 2, 3) 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. Five days following the first topical application of the test item or vehicle (on Day 6) all mice were injected via the tail vein with 250 µL of phosphate buffered saline (PBS) containing ³HTdR at 80 µCi/mL (specific activity 2.0 Ci/mmoL, ARC UK Ltd), giving a total of 20 µCi to each mouse. Five hours later, draining auricular lymph nodes were excised and pooled for each experimental group. A single cell suspension of pooled lymph node cells was prepared by gentle mechanical disaggregation through 200-mesh stainless steel gauze. The lymph node cells per group were each rinsed through the gauze with 4 mL of PBS into a petri dish and then transferred to the respective centrifuge tube. The petri dish was washed with an additional 5 mL of PBS to remove all remaining lymph node cells and these were added to the centrifuge tube. The pooled lymph node cells were pelleted by centrifugation and the obtained pellet was resuspended in 10 mL of PBS and re-pelleted. Then, the pellet was resuspended in 3 mL of 5% trichloroacetic acid (TCA) and precipitated at 4 °C for a period of approx. 18 h. After centrifugation, the pellet was resuspended in 5% TCA and ³HTdR incorporation was measured by β-scintillation counting.

Positive control substance(s):

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

Statistics:

The mean ear thickness was determined in control and treated animals.

Positive control results:

The current positive control substance α-hexylcinnamaldehyde at 25% v/v in acetone/olive oil 4:1 produced a stimulation index (SI) of 5.76, thus fulfilling the reliability criteria for the LLNA (SI > 3).

Key result

Parameter:

SI

Value:

9.2

Test group / Remarks:

25% in AOO 4:1

Key result

Parameter:

SI

Value:

12.84

Test group / Remarks:

50% in AOO 4:1

Key result

Parameter:

SI

Value:

11.55

Test group / Remarks:

100%

Table 1. Results of the LLNA

Concentration (% v/v) in acetone/olive oil 4:1	dpm	dpm/Node^a	Stimulation Index^b	Result
Vehicle	12918.23	1614.78	na	na
25	11883.60	14860.45	9.20	positive
50	165885.80	20735.73	12.84	positive
100	149168.70	18646.09	11.55	positive

dpm = Disintegrations per minute

a = Disintegrations per minute/node obtained by dividing the disintegrations per minute value by 8 (total number of lymph nodes)

b = Stimulation Index of 3.0 or greater indicates a positive result

na = Not applicable

CLINICAL OBSERVATIONS

No mortalities and no signs of systemic toxicity were observed during the study.

 

BODYWEIGHTS

Bodyweight changes of the test animals between Day 1 and Day 6 were comparable to those observed in the corresponding control group animals over the same period.

Interpretation of results:

Category 1 (skin sensitising) based on GHS criteria

Conclusions:

CLP: Skin Sens. 1, H317

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

A Local Lymph Node Assay with 2-methoxyethyl acrylate was performed in female mice according to OECD 429 and in compliance with GLP (Henzell, 2012). Based on the results of a preliminary range finding test, 5 animals per group were epidermally treated (25 µL/ear) with the test substance at concentrations of 25, 50 and 100% in acetone/olive oil (4:1 v/v) for 3 consecutive days. A similar constituted group was treated with the vehicle alone and served as controls. On Day 6, all mice were injected with ³H-methyl thymidine (³HTdR) and sacrificed 5 h afterwards. The draining auricular lymph nodes were excised and pooled for each experimental group. ³HTdR incorporation in lymphocytes was measured by beta-scintillation counting. There were no deaths during the study period and no signs of toxicity or local skin irritation were noted. The mean disintegration per minute (DPM) values for the single cell suspensions of each experimental group were 14860.45, 20735.73 and 18646.09 at concentrations of 25, 50 and 100% of the test substance in acetone/olive oil (4:1 v/v), respectively, and thus significantly increased compared to values of the control value (DPM = 1614.78). The stimulation indices were 9.20, 12.84 and 11.55 for concentrations of 25, 50 and 100%, respectively. No EC3 value was established, since no conventional dose-response relationship was observed. Since all SI values were greater than 3, the substance was considered to be a skin sensitiser.

 

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Harmonised classification

2-methoxyethyl acrylate has been inserted into ATP15. The following classification applies:

• Skin Sens. 1 – H317: May cause and allergic skin reaction

 

Details:

Skin sensitisation

The LLNA performed on CBA/Ca Mice with 2-methoxyethyl acrylate was positive, with a stimulation index higher than 3 (SI >9.2% and above) at the concentrations of 25%, 50% and 100%. The EC3 value is not available and there is no experimental data with which to calculate it.

Since all tested concentrations were above 2%, there are no data to exclude the possibility that at lower concentrations (≤ 2%) 2-methoxyethyl acrylate does not stimulate cell proliferation with a stimulation index above 3, therefore it is not possible to exclude that the substance meets classification criteria for category 1A. It is not known if the result is positive at a concentration of ≤2%, therefore it is not possible to assign a subcategory.

Therefore, 2-methoxyethyl acrylate is classified Skin Sens. 1; H317 “May cause an allergic skin reaction” with no subcategorization.

 

Respiratory sensitisation

No specific animal or human data was available on 2-methoxyethyl acrylate.

According to the OECD QSAR Toolbox, version 3.4., acrylates have been suggested to be capable of reacting with proteins in the lung via a direct Michael addition mechanism. The Leadscope Toxicity Database also predicts positive results for this substance. DEREK nexus does not predict respiratory sensitisation for 2-methoxyethyl acrylate as no structural alerts for acrylates were developed in the model.

With regard to the predicted metabolites (WHO, 2009) only formaldehyde (a known metabolite of 2-methoxyethanol) has a harmonised classification for respiratory sensitisation. Respiratory sensitisation has not been reported for the expected primary metabolites of 2-methoxyethyl acrylate, 2-methoxyethanol and acrylic acid.

No human or animal data are available specifically on 2-methoxyethyl acrylate on respiratory sensitisation.

RAC is of the opinion that the available data are insufficient to classify 2-methoxyethyl acrylate as a respiratory sensitizer in agreement with the DS.