2,3-Epoxypropyl neodecanoate, oligomeric reaction products with cyclohexane-1,2-dicarboxylic anhydride and propylidenetrimethanol

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 June - 26 July 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

July 2010

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

May 2008

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Version / remarks:

March 2003

Deviations:

no

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA): DA

Test material

Test material information

Constituent 1

Reference substance name:

2,3-Epoxypropyl neodecanoate, oligomeric reaction products with cyclohexane-1,2-dicarboxylic anhydride and propylidenetrimethanol

EC Number:

500-334-1

EC Name:

2,3-Epoxypropyl neodecanoate, oligomeric reaction products with cyclohexane-1,2-dicarboxylic anhydride and propylidenetrimethanol

Cas Number:

154565-28-3

Molecular formula:

A molecular formaula is not available for this UVCB

IUPAC Name:

2,3-Epoxypropyl neodecanoate, oligomeric reaction products with cyclohexane-1,2-dicarboxylic anhydride and propylidenetrimethanol

Test material form:

solid

Remarks:

Clear yellow resin

Specific details on test material used for the study:

Identification: Epoxypropyl neodecanoate, oligomeric reaction products with cyclohexane-1,2-dicarboxylic anhydride and propylidenetrimethanol
Appearance: Clear yellow resin
Batch: CVR 83297
Storage: At room temperature protected from light
Stable under storage conditions until: 27 December 2018 (expiry date)

In vivo test system

Test animals

Species:

mouse

Strain:

CBA:J

Sex:

female

Details on test animals and environmental conditions:

Species: Mouse
Strain: CBA/J
Condition: Inbred, SPF-Quality
Source: Janvier, Le Genest-Saint-Isle, France
Number of Animals: 20 Females (nulliparous and non-pregnant). Five females per group.
Age at the Initiation of Dosing: Young adult animals (approximately 8 weeks old) were selected.
Weight at the Initiation of Dosing: 18.9-22.6 g.

On arrival and following assignment to the study, animals were group housed (up to 5 of the same dosing group together) in polycarbonate cages (Makrolon) containing sterilized sawdust as bedding material. Target temperatures of 18 to 24°C with a relative target humidity of 40-70% were maintained. The actual daily mean temperature during the study period was 22°C with an actual daily mean relative humidity of 47-68%. A 12-hour light/12-hour dark cycle was maintained. Ten or greater air changes per hour with 100% fresh air (no air recirculation) were maintained in the animal rooms.

Pelleted diet and drinking water were provided ad libitum throughout the study.

Study design: in vivo (LLNA)

Vehicle:

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

Concentration:

10%, 25%, 50%

No. of animals per dose:

5 female

Details on study design:

A pre-screen was conducted in order to select the highest test item concentration to be used in the main study. In principle, this highest concentration should cause no systemic toxicity, may give well-defined irritation as the most pronounced response (maximum grade 2 and/or an increase in ear thickness < 25%) and/or is the highest possible concentration that can technically be applied. Two test item concentrations were tested; a 25% and 50% concentration. The highest concentration was the maximum concentration required in the test guidelines.

In the main study, three groups of five animals were treated with one concentration (10%, 25% or 50%). The highest concentration was selected from the pre-screen test. One group of five animals was treated with the vehicle. The dorsal surface of both ears was topically treated (25 ¿L/ear) at approximately the same time on each day (Days 1, 2, 3). The formulations were stirred with a magnetic stirrer immediately prior to dosing. The control animals were treated in the same way as the experimental animals, except that the vehicle was administered instead of the test material.

On Day 6, each animal was injected via the tail vein with 0.25 mL sterile phosphate buffered saline (PBS) containing 20 ¿Ci 3H-methyl thymidine. After five hours, all animals were killed by intraperitoneal injection (0.2 mL/animal) of Euthasol®. The draining (auricular) lymph node of each ear was excised. The relative size of the nodes was estimated by visual examination and abnormalities of the nodes and surrounding area were recorded. The nodes were pooled for each animal in PBS. Following excision of the nodes, a single cell suspension of lymph node cells was prepared in PBS by gentle separation through stainless steel gauze (200 ¿m). Cells were washed twice with an excess of PBS by centrifugation at 200g for 10 minutes at 4ºC. To precipitate DNA, cells were exposed to 5% trichloroacetic acid and stored overnight. Precipitates were recovered by centrifugation, resuspended in 1 mL TCA and radioactivity measured by LSC.

Positive control substance(s):

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

Statistics:

Not required

Results and discussion

Positive control results:

The results of a separate reliability check (Test Facility Study No. 518176) are reported. The check was performed in June 2017 using 5%, 10% and 25% hexyl cinnamic aldehyde. SI values of 1.2, 2.3 and 5.6 are reported (EC3: 13.2%), confirming the sensitivity of the assay.

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

1.4

Variability:

standard deviation 0.4

Test group / Remarks:

10%

Results 2

Key result

Parameter:

SI

Value:

1.4

Variability:

standard deviation 0.3

Test group / Remarks:

25%

Results 3

Key result

Parameter:

SI

Value:

2.6

Variability:

standard deviation 0.5

Test group / Remarks:

50%

Results 4

Key result

Parameter:

EC3

Value:

> 50

Remarks on result:

other: EC3 could not be calculated as all SI values are <3

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
Mean dpm/animal values for the experimental groups treated with concentrations of 10, 25 and 50% were 566, 583 and 1076, respectively. The mean dpm/animal value for the vehicle control group was 414. The SI values calculated for concentrations of 10, 25 and 50% were 1.4, 1.4 and 2.6, respectively.

EC3 CALCULATION
An EC3 could not be calculated (>50%).

CLINICAL OBSERVATIONS:
In the pre-screen, very slight irritation of the ears was shown by animals treated at 50% at Days 3 and 4, no further erythema was noted. Variations in ear thickness during the observation period were less than 25%. In the main study, no mortality occurred and no clinical signs of systemic toxicity were observed in the animals of the main study. Alopecia between the ears and neck was noted for all animals treated at 50% between Days 4 and 6, which was considered not to have a toxicologically significant effect on the activity of the nodes. In the main study, very slight or well defined erythema of the ears was shown by several animals treated at 25% and all animals treated at 50% between Days 2 and 5 but was considered not to have a toxicologically significant effect on the activity of the nodes.

BODY WEIGHTS:
Body weights and body weight gain of experimental animals remained in the same range as controls over the study period.

Any other information on results incl. tables

The majority of auricular lymph nodes were considered normal in size, except for the nodes in one animal treated at 25% and all animals treated at 50%, which were considered enlarged. No macroscopic abnormalities of the surrounding area were noted for any of the animals.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

As all SI values were <3, the EC3 was calculated to be >3. CLP criteria for classification are not met.

Executive summary:

The objective of this study was to evaluate whether the test material [Epoxypropyl neodecanoate, oligomeric reaction products with cyclohexane-1,2-dicarboxylic anhydride and propylidenetrimethanol] shows skin sensitisation potential in mice after three epidermal exposures (LLNA, OECD 429). Test material concentrations selected for the main study were based on the results of a pre-test. In the main study, three experimental groups of five female CBA/J mice were treated with concentrations of 10, 25 or 50% w/w on three consecutive days, by open application on the ears. Five vehicle control animals were similarly treated, but with the vehicle alone (acetone/olive oil (4:1 v/v)). Three days after the last exposure, all animals were injected with 3H-methyl thymidine and after five hours the draining (auricular) lymph nodes were excised and pooled for each animal. After precipitating the DNA of the lymph node cells, radioactivity measurements were performed. Activity was expressed as the number of disintegrations per minute (dpm) and a stimulation index (SI) was subsequently calculated for each group. The majority of auricular lymph nodes were considered normal in size, except for the nodes in some of the animals treated at 25% and all animals treated at 50%, which appeared larger in size as compared to the other treated groups. No macroscopic abnormalities of the surrounding area were noted for any of the animals. Mean dpm values for the experimental groups treated with concentrations of 10, 25 and 50% were 566, 583 and 1076, respectively. The mean dpm value for the vehicle control group was 414. SI values calculated for the test material concentrations of 10%, 25% and 50% were therefore 1.4, 1.4 and 2.6, respectively. As all of the SI values were <3, the test material was not considered to have skin sensitisation potential. The EC3 value exceeds 50%. A six-month reliability check with alpha-hexylcinnamaldehyde confirmed the sensitivity of the assay.