2-Propenoic acid, 2-methyl-, monoester with 1,2-propanediol, reaction products with dipropylene glycol and 1,1'-methylenebis[isocyanatobenzene]

Skin sensitisation

Administrative data

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: GLP-guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted in 2010

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

(Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, Landesinstitut für Arbeitsschutz und Produktsicherheit, München, Germany)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

UMA121

IUPAC Name:

UMA121

Constituent 2

Reference substance name:

Propanol, oxybis-, oligomeric reaction products with 1,1'-methylenebis[isocyanatobenzene], propylene glycol monomethacrylate-blocked

EC Number:

923-201-3

Cas Number:

1192143-92-2

Molecular formula:

C29H34N2O8 C50H58N4O13 C71H82N6O18 C92H106N8O23 C113H130N10O28

IUPAC Name:

Propanol, oxybis-, oligomeric reaction products with 1,1'-methylenebis[isocyanatobenzene], propylene glycol monomethacrylate-blocked

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/CaOlaHsD

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Winkelmann, Borchen, Germany
- Age at study initiation: 8 - 9 weeks
- Weight at study initiation: 19 - 22 g
- Housing: The animals were housed in groups of 3 (pre-screen test) and 5 (main study) in IVC cages, type II L, polysulphone cages on Altromin saw fibre bedding.
- Diet: Altromin 1324 maintenance diet for rats and mice, ad libitum
- Water: tap water, sulphur acidified to a pH value of approximately 2.8, ad libitum
- Acclimation period: at least 5 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 10
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

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

Concentration:

12.5, 25 and 50% (w/v)

No. of animals per dose:

5

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: The maximum technically applicable concentration of the test substance was found to be 50% in acetone/olive oil (4:1) (AOO).
- Irritation: Two animals were treated with 50% test substance in AOO. A third animals was treated with vehicle only and served as negative control. No signs of systemic toxicity or skin irritation (ear swelling, erythema and eschar formation) were detected in the test substance-treated animals. However, alopecia was observed at the apllication site.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: ³H-Methylthymidine incorporation determined by β-scintillation.
- Criteria used to consider a positive response: The substance is regarded as skin sensitiser if at least one concentration of the test substance results in a 3-fold or greater increase in ³H-Methylthymidine incorporation into lymph node cells of the test group animals, relative to that recorded for the lymph nodes of the control group (SI ≥ 3.0).

TREATMENT PREPARATION AND ADMINISTRATION: Each mouse was treated by topical application of 25 µL of a test substance solution or the vehicle alone to the dorsal surface of each ear. Topical applications were performed once daily over three consecutive days.
Five days after the first topical application, all mice were dosed with 20 µCi ³H-Methylthymidine by intravenous injection of 250 µL ³H-Methylthymidine into the tail vein. Approximately 5 hours after the injection, all mice were sacrificed by cervical dislocation. The draining auricular lymph nodes were excised, individually pooled for each animal (2 lymph nodes per animal) and collected in phosphate buffered saline (PBS). A single cell suspension of pooled lymph node cells was prepared by gentle mechanical disaggregation through polyamide gauze (200 mesh size). The cell suspension was washed with PBS, pelleted and resuspended in 1 mL 5% TCA and 7 mL scintillation fluid was added. The ³H-Methylthymidine incorporation was measured in a β-counter and expressed as the number of disintegrations per minute (DPM) individually for each animal.

Positive control substance(s):

other: 1% p-phenylenediamine (reliability check)

Statistics:

Outlier tests for statistical significance were performed according to Grubbs, Nalimov and Dixon.

Results and discussion

Positive control results:

A reliability test with the positive control substance was routinely performed 2 months prior to the present study. A mean SI value of 11.3 ± 1.7 (n = 5) was determined.

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: 12.5% test substance: 1.4 25% test substance: 1.5 50% test substance: 1.9 negative control: 1.0

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: 12.5% test substance: 1902.4 25% test substance: 2022.8 50% test substance: 2505.8 negative control: 1346.2

Any other information on results incl. tables

All animals survived until the end of the study period without showing any clinical signs or signs of dermal irritation at the application site. Alopecia was observed at the application site from Day 3 until the end of the study period in all animals treated with the test substance at a concentration of 50%. The treatment with the test substance did not influence the body weight gain of the treated animals.

Table 1. Radioactive determination of ³H-Methylthymidine incorporation

	Animal No.	CPM	DPM	DPM - background	SI
12.5% test substance	1	789	1626	1573.8	1.2
	2	1107	2283	2230.8	1.7
	3	827	1721	1668.8	1.3
	4	1102	2294	2241.8	1.7
	5	772	1588	1535.8	1.2
	mean	919.4	1902.4	1850.2	1.4
	SD	152.2	318.2	318.2	0.2
25% test substance	6	1033	2145	2092.8	1.6
	7	837	1738	1685.8	1.3
	8	2705*	5643*	---	---
	9	1241	2538	2485.8	1.9
	10	812	1670	1617.8	1.3
	mean	980.8	2022.8	1970.6	1.5
	SD	172.9	348.5	348.5	0.3
50% test substance	11	1015	2096	2043.8	1.6
	12	1298	2679	2626.8	2.0
	13	1275	2686	2633.8	2.0
	14	1820**	3777**	---	---
	15	1241	2562	2509.8	1.9
	mean	1207.3	2505.8	2453.6	1.9
	SD	112.8	241.6	241.6	0.2
negative control	16	243	497	444.8	---
	17	479	988	935.8	---
	18	667	1375	1322.8	---
	19	981	2014	1961.8	---
	20	903	1857	1804.8	---
	mean	654.6	1346.2	1294.0	1.0
	SD	271.5	558.1	558.1	---
Background value	mean	25.8	52.2	---	---
	SD	1.5	3.5	---	---

* outlier, failed Grubbs, Nalimov and Dixon

** outlier, failed Nalimov and Dixon

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified