Isodecyl methacrylate

Endpoint summary

Description of key information

Isodecyl methacrylate was demonstrated to be not sensitising in a fully valid GLP LLNA guideline study according to OECD 429.

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:

2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Specific details on test material used for the study:

- Name of test material (as cited in study report): Isodecyl methacrylate (CAS 29964-84-9)

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Netherlands
B.V. Postbus 6174
5960 AD Horst / The Netherlands
- Age at study initiation: 8 - 12 weeks (beginning of treatment)
- Weight at study initiation: 19.3 - 21.9 g (mean)
- Housing: single
- Diet (e.g. ad libitum): Pelleted standard diet, ad libitum
(Harlan Laboratories B.V., 5960 AD Horst / Netherlands)
- Water (e.g. ad libitum): Tap water, ad libitum
- Acclimation period: At least 5 days prior to the start of dosing under test conditions after health examination. Only animals without any visible signs of illness were used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Temperature 22 +- 2°C
- Humidity (%): Relative humidity 45 - 65%
- Photoperiod (hrs dark / hrs light): Artificial light 6.00 a.m. - 6.00 p.m.

Vehicle:

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

Concentration:

25 %, 50 % and 100 %

No. of animals per dose:

4

Details on study design:

In order to study a possible allergenic potential of Isodecyl methacrylate, three groups each of four female mice were treated with different concentrations of the test item by topical application at the dorsum of each ear lobe (left and right) on three consecutive days. A control group of four mice was treated with the vehicle only. Five days after the first topical application, the mice were intravenously injected into a tail vein with radio-labelled thymidine (3H-methyl thymidine). Approximately five hours after intravenous injection, the mice were sacrificed and the draining auricular lymph nodes excised and pooled per group. Single cell suspensions of lymph node cells were prepared from pooled lymph nodes, which were subsequently washed and incubated with trichloroacetic acid overnight. The proliferative capacity of pooled lymph node cells was determined by the incorporation of 3H-methyl thymidine measured in a -scintillation counter.



Experimental Design and Procedures
Topical Application
Each test group of mice was treated by topical (epidermal) application to the dorsal surface of each ear lobe (left and right) with different test item concentrations of 25, 50, and 100% (w/v) in acetone:olive oil (4+1). The application volume, 25 µl, was spread over the entire dorsal surface (  8 mm) of each ear lobe once daily for three consecutive days. A further group of mice was treated with an equivalent volume of the relevant vehicle alone (control animals).
Administration of 3H-Methyl Thymidine
3H-methyl thymidine (3HTdR) was purchased from GE Healthcare (GE Healthcare product code no. TRA 310; specific activity, 2 Ci/mmol; concentration, 1 mCi/ml).
Five days after the first topical application, all mice were administered with 250 µl of 78.3 µCi/ml 3HTdR (corresponds to 19.6 µCi 3HTdR per mouse) by intravenous injection via a tail vein.
Determination of Incorporated 3HTdR
Approximately five hours after treatment with 3HTdR all mice were euthanised by intraperitoneal injection of Pentobarbital-Natrium (Release, WDT, D-30827 Garbsen).
The draining lymph nodes were rapidly excised and pooled per group (8 nodes per group). Single cell suspensions (in phosphate buffered saline) of pooled lymph node cells were prepared by gentle mechanical disaggregation through stainless steel gauze (200 µm mesh size). After washing two times with phosphate buffered saline (approx. 10 ml) the lymph node cells were resuspended in 5 % trichloroacetic acid (approx. 3 ml) and incubated at approximately +4 °C for at least 18 hours for precipitation of macromolecules. The precipitates were then resuspended in 5 % trichloroacetic acid (1 ml) and transferred to plastic scintillation vials with 10 ml of ‘Ultima Gold’ scintillation liquid (Perkin Elmer (LAS) GmbH, D-63110 Rodgau) and thoroughly mixed.
The level of 3HTdR incorporation was then measured on a -scintillation counter (Tricarb 2900 TR, Perkin Elmer (LAS) GmbH, D-63110 Rodgau). Similarly, background 3HTdR levels were also measured in two 1ml-aliquots of 5 % trichloroacetic acid. The -scintillation counter expresses 3HTdR incorporation as the number of radioactive disintegrations per minute (DPM).

Interpretation of Raw Data
The proliferative response of lymph node cells is expressed as the number of radioactive disintegrations per minute per lymph node (DPM/node) and as the ratio of 3HTdR incorporated into lymph node cells of test lymph nodes relative to that recorded for control lymph nodes (stimulation index). Before DPM/node values were determined, mean scintillation-background DPM was subtracted from test and control raw data.
A test item is regarded as a sensitiser in the LLNA if the following criteria are fulfilled:
- First, that exposure to at least one concentration of the test item resulted in an incorporation of 3HTdR at least 3-fold or greater than that recorded in control mice, as indicated by the stimulation index.
- Second, that the data are compatible with a conventional dose response, although allowance must be made (especially at high topical concentrations) for either local toxicity or immunological suppression.
Observations
In addition to the sensitising reactions the following observations and data were recorded during the test and observation period:
Mortality / Viability once daily (week day) from experimental start to necropsy.
Body weights prior to the first application and prior to treatment with 3HTdR.
Clinical signs (local / systemic) In the pre-test clinical signs were recorded within 1 hour and 24 ± 4 hours after each application as well as on day 7. In the main experiment clinical signs were recorded within 1 hour each application, and 24 ± 4 hours after the first and second application as well as on the day of preparation. Especially the treatment sites were observed carefully.

Positive control substance(s):

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

Statistics:

The mean values and standard deviations were calculated in the body weight tables.

Positive control results:

Results of the GLP Positive Control

Experiment performed in June 2009.
Positive control substance: alpha-Hexylcinnamaldehyde
Vehicle: acetone:olive oil (4+1)
Test item concentration % (w/v)

Group Measurement DPM Calculation Result
DPM-BG a) number of lymph nodes DPM per lymph node b) S.I.
--- BG I 23 --- --- --- ---
--- BG II 19 --- --- --- ---
0 1 5842 5821 8 727.6
5 2 10450 10459 8 1303.6 1.79
10 3 12168 12147 8 1518.4 2.09
25 4 39834 39813 8 4976.6 6.84

BG = Background (1 ml 5% trichloroacetic acid) in duplicate
1 = Control Group
2-4 = Test Group
S.I. = Stimulation Index
a) = The mean value was taken from the figures BG I and BG II
b) = Since the lymph nodes of the animals of a dose group were pooled, DPM/node was determined by dividing the
measured value by the number of lymph nodes pooled

Test item concentration % S.I.
Test Group 3 10 (a) 2.09 (b)
Test Group 4 25 (c) 6.84 (d)

EC3 = (a-c) [(3-d)/(b-d)] + c = 12.9 % (w/v)
EC3 = Estimated concentration for a S.I. of 3.
a,b,c,d = Co-ordinates of the two pairs of data lying immediately above and below the S.I. value of 3 on the LLNA dose response plot.

Parameter:

SI

Value:

1.43

Test group / Remarks:

25% / number of lymph nodes: 8

Remarks on result:

other: see Remark

Remarks:

In this study Stimulation Indices (S.I.) of 1.43, 1.22 and 0.97 were determined with the test item at concentrations of 25, 50, and 100% in acetone : olive oil (4+1), respectively. The EC3 value could not be calculated, since all induced S.I.'s are below 3.

Parameter:

SI

Value:

1.22

Test group / Remarks:

50% / number of lymph nodes: 8

Remarks on result:

other:

Remarks:

In this study Stimulation Indices (S.I.) of 1.43, 1.22 and 0.97 were determined with the test item at concentrations of 25, 50, and 100% in acetone : olive oil (4+1), respectively. The EC3 value could not be calculated, since all induced S.I.'s are below 3.

Parameter:

SI

Value:

0.97

Test group / Remarks:

100% / number of lymph nodes: 8

Remarks on result:

other:

Remarks:

In this study Stimulation Indices (S.I.) of 1.43, 1.22 and 0.97 were determined with the test item at concentrations of 25, 50, and 100% in acetone : olive oil (4+1), respectively. The EC3 value could not be calculated, since all induced S.I.'s are below 3.

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: see below

Calculation and Results of Individual Data

 

Vehicle: acetone : olive oil (4 +1)

Test item concentration % (w/v)	Group	Measurement DPM	Calculation			Result
			DPM-BG^a)	number of lymph nodes	DPM per lymph node^b)	S.I.
---	BG I	21	---	---	---	---
---	BG II	31	---	---	---	---
---	CG1	4282	4256	8	523.0
25	2	6106	6080	8	760.0	1.43
50	3	5235	5209	8	651.1	1.22
100	4	4162	4136	8	517.0	0.97

 

BG  =   Background (1 ml 5% trichloroacetic acid) in duplicate

CG1=   Control Group

2-4 =   Test Group

S.I.  =   Stimulation Index

^a)    =   The mean value was taken from the figures BG I and BG II

^b)     =   Since the lymph nodes of the animals of a dose group were pooled, DPM/node was determined by dividing the measured value by the number of lymph nodes pooled

The EC3 value could not be calculated, since all SI´s are below 3.

 

Viability / Mortality

No deaths occurred during the study period.

Clinical Signs

No symptoms of local toxicity at the ears of the animals and no systemic findings were observed during the study period

Body Weights

The body weight of the animals, recordedprior to the first application and prior to treatment with³HTdR, was within the range commonly recorded for animals of this strain and age.

The individual body weight values are included in the following table:

 

Tables of Body Weights

 

Animal No.	Dose Group	Initial Weight (g)	weight prior to treatment with^3HTdR (g)
1	1	21.1	21.5
2	1	20.0	20.9
3	1	19.7	20.1
4	1	21.9	22.6
5	2	21.5	23.2
6	2	20.8	22.3
7	2	21.1	21.4
8	2	21.6	23.5
9	3	19.7	20.6
10	3	19.7	21.1
11	3	21.0	21.4
12	3	20.2	20.0
13	4	19.8	21.3
14	4	20.7	21.4
15	4	19.3	20.0
16	4	20.8	20.2
	Mean	20.6	21.3
	Standard Deviation	0.8	1.1

 

Interpretation of results:

GHS criteria not met

Conclusions:

Isodecyl methacrylate was found to be not a skin sensitiser under the described conditions.

Executive summary:

In order to study a possible contact allergenic potential of Isodecyl methacrylate, three groups each of four female mice were treated daily with the test item at concentrations of 25, 50, and 100% (w/v) in acetone:olive oil (4+1) by topical application to the dorsum of each ear lobe (left and right) for three consecutive days. A control group of four mice was treated with the vehicle (acetone:olive oil (4+1)) only. Five days after the first topical application the mice were injected intravenously into a tail vein with radio-labelled thymidine (³H-methyl thymidine). Approximately five hours after intravenous injection, the mice were sacrificed, the draining auricular lymph nodes excised and pooled per group. Single cell suspensions of lymph node cells were prepared from pooled lymph nodes, which were subsequently washed and incubated with trichloroacetic acid overnight. The proliferative capacity of pooled lymph node cells was determined by the incorporation of³H-methyl thymidine measured in ab-scintillation counter. All treated animals survived the scheduled study period and no signs of toxicity were observed. A test item is regarded as a sensitiser in the LLNA if the exposure to one or more test concentration resulted in 3-fold or greater increase in incorporation of³HTdR compared with concurrent controls, as indicated by the Stimulation Index (S.I.). The estimated concentration of test item required to produce a S.I. of 3 is referred to as the EC3 value.In this study Stimulation Indices of 1.43, 1.22, and 0.97 were determined with the test item at concentrations of 25, 50, and 100% in acetone:olive oil (4+1). The EC3 value could not be calculated, since none of the tested concentrations induced an S.I. greater than 3.

In conclusion, Isodecyl methacrylate was not a skin sensitiser when tested in the Local Lymph Node Assay according to OECD TG 429.

NOTE: Any of data in this dataset are disseminated by the European Union on a right-to-know basis and this is not a publication in the same sense as a book or an article in a journal. The right of ownership in any part of this information is reserved by the data owner(s). The use of this information for any other, e.g. commercial purpose is strictly reserved to the data owners and those persons or legal entities having paid the respective access fee for the intended purpose.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Isodecyl methacrylate was demonstrated to be not sensitising in a fully valid GLP in vivo LLNA guideline study according to OECD 429.

In vivo dermal sensitization results, weight of evidence (WoE) determinations, and results for EpiSensA and other in silico, in chemico, and in vitro sensitization assays for IDMA

	In vivo LLNA		EpiSensA
Test chemical	WoE hazard	WOE potency	Hazard	Potency	h-CLAT	TIMES-SS	DPRA	LuSens (L)/KeratinoSens (K)
IDMA	nonsensitizer	nonsensitizer	positive	weak	positive	weak positive	negative	negative (K)

Abbreviations: DPRA, direct peptide reactivity assay; EpiSensA, epidermal sensitization assay; h-CLAT, human cell line activation test; TIME-SS, times metabolism simulator platform for predicting skin sensitization.

Kimber, I. (2019). The activity of methacrylate esters in skin sensitisation test methods: A review. Regulatory Toxicology and Pharmacology, 104, 14–20. https://doi.org/10.1016/j.yrtph.2019.02.014

cited from: Mizumachi H, LeBaron MJ, Settivari RS, Miyazawa M, Marty MS, Sakaguchi H. Characterization of dermal sensitization potential for industrial or agricultural chemicals with EpiSensA. J Appl Toxicol. 2020;1–13. https://doi.org/10.1002/jat.4076

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

According to the criteria as of directive 1272/2008/EC, no classification is warranted.