Reaction mass of (2,4,6-trioxo-1,3,5-triazine-1,3,5(2H,4H,6H)-triyl)tri-2,1-ethanediyl triacrylate and 2-Propenoic acid, 1,1'-[[dihydro-5-(2-hydroxyethyl)-2,4,6-trioxo-1,3,5-triazine-1,3(2H,4H)-diyl]di-2,1-ethanediyl] ester

Dermal absorption

Administrative data

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study performed in accordance to OECD 428 without any deviations to study plan (See attached study report)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 428 (Skin Absorption: In Vitro Method)

Deviations:

no

Principles of method if other than guideline:

NA

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

2-ethyl-2-[[(1-oxoallyl)oxy]methyl]-1,3-propanediyl diacrylate

EC Number:

239-701-3

EC Name:

2-ethyl-2-[[(1-oxoallyl)oxy]methyl]-1,3-propanediyl diacrylate

Cas Number:

15625-89-5

Molecular formula:

C15H20O6

IUPAC Name:

2,2-bis(prop-2-enoyloxymethyl)butyl prop-2-enoate

Test material form:

solid - liquid: suspension

Details on test material:

Non-radiolabelled TMPTA:
- Name of test material (as cited in study report): TMPTA
- Physical state: Clear, colorless liquid
- Analytical purity: 80.2 %
- Lot/batch No.: 41095133SP01
- Expiration date of the lot/batch: 30 September 2015
- Storage condition of test material: At room temperature protected from light

Radiolabelled TMPTA:
- Name of test material (as cited in study report): [14C]TMPTA
- Molecular weight: 296.35 g.mol-1 (at this specific activity)
- Specific activity: 1 mCi/mmol (0.125 MBq/mg)
- Radiochemical purity: 97.3%
- Lot/batch No.: 361-119-001-A-20150804-DRE
- Date of arrival at TNO Triskelion : 17 August 2015
- Expiration date of the lot/batch: 17 August 2020
- Storage condition of test material: < –18 ºCt
N.B. Due to the product being subject to polymerization, the material is shipped copresent with a stabilizer. The purity of the tracer was checked prior to the conduct of the study and was found to be > 96 %.

Radiolabeled reference substance:
- Name : [3H]H2O
- Batch number: 1773956
- Specific activity: 37 MBq.g-1
- Radiochemical purity : not determined
- Molecular weight: 18.0 g.mol-1
- Date of arrival at TNO Triskelion : 13 May 2014
- Expiry date: 13 May 2019 (default, assigned by TNO Triskelion)
- Supplier: PerkinElmer Life Sciences Inc.
- Storage conditions: 2 – 10 ºC
- TNO Triskelion reference no. : TT132 (radioactive materials)

Radiolabelling:

yes

Remarks:

[14C]TMPTA

Test animals

Species:

other: In vitro

Strain:

other: human skin samples

Sex:

not specified

Details on test animals or test system and environmental conditions:

Preparation of skin membranes:
Human skin membranes were prepared from frozen skin samples, present at TNO Triskelion. Human skin, all derived from the breast, was obtained from four female donors directly after surgery.
Donor 1: H15/05, born in 1966, arrival on 15 January 20151
Donor 2: H15/25, born in 1985, arrival on 7 April 20151
Donor 3: H15/26, born in 1969, arrival on 7 April 2015
Donor 4: H15/35, born in 1952, arrival on 26 June 2015

Administration / exposure

Duration of exposure:

TMPTA was topically applied to the skin membranes. The exposure time was 8h and receptor fluid samples were collected from 0-24 h.

Doses:

Two skin membranes from four donors in each test group was used (a total of 8).

Total concentration measured was 910 g/L. Mean dose applied was 9098 +/- 108 ug/cm2 (a net volume of ca. 6.4 μL of the dose preparations was applied on each skin membrane (0.64 cm2).

The dose preparations were prepared within one day prior to application as follows:
- Amount of [14C] TMPTA was 586 uL (~1.3 MBq, ~10.4 mg)
- Non-radiolabelled TMPTA was 0.5644 g
- Total concentration measured was 910 g/L
- Radioactive concentration measured was 2.49 MBq/mL

No. of animals per group:

NA

Control animals:

no

Remarks:

NA

Details on study design:

NA

Details on in vitro test system (if applicable):

SKIN PREPARATION
Human skin, all derived from the breast, was obtained from four female donors directly after surgery from hospital. The transportation of the skin to the laboratory was carried out as soon as possible after dissection, while the skin was placed in a plastic container that was kept on ice. Subcutaneous fat was removed and the skin was stored at < −18 °C until use (For logistic reasons, the skin was kept in the refrigerator (2-10 ºC) overnight or over the weekend. Prolonged storage at 2-10 ºC is considered not to have affected the quality of the skin tissue, which was confirmed by the
data from the integrity test). In agreement with the hospital, only skin tissue for which informed consent was given by the donor, was provided to TNO Triskelion. Upon thawing, the skin was cut to a target thickness of ca. 0.2-0.4 mm (i.e. splitthickness skin membranes) using a Dermatome (25 mm, Nouvag GmbH, Germany). The thickness of all skin preparations was measured with a digimatic micrometer (Mitutoyo Corporation, Japan).

PRINCIPLES OF ASSAY
Flow-through diffusion cells and receptor fluid:
Approximately 20 h before the start of exposure to the test preparation, the split thickness skin membranes were placed in 9 mm flow-through automated diffusion cells (PermeGear Inc., Riegelsville, PA, USA) to hydrate the skin. The mean skin surface temperature was 32 ± 1 ºC and exposure was at ambient humidity. Following application of the test preparations, the actual temperature was recorded at 15-minute intervals during the study in one diffusion cell containing a non-exposed skin membrane. The receptor fluid was pumped at a flow rate of ca. 1.8 mL.h-1 and consisted of phosphate buffered saline (PBS, containing 0.01% sodium azide, w/v), supplemented with 6% polyoxyethylene 20-oleyl ether (w/v),pH 7.2.

After placing the skin membranes in the diffusion cells, membrane integrity was assessed. To this purpose, the permeability coefficient (Kp) for tritiated water was determined as follows: 200 μL saline (supplemented with 0.01% sodium azide) containing [3H]H2O (17.2 kBq.mL-1) was applied in the donor compartment of the flowthrough diffusion cells. The compartments were covered with a glass slide. Samples of the receptor fluid (ca. 1.8 mL per hour) were collected every hour up to three hours after application. Tritiated water remaining at the application site was then removed and the skin was dried with cotton swabs. Membranes were kept overnight to allow wash-out of the tritiated water from the skin.

Results and discussion

Absorption in different matrices:

See below

Total recovery:

See below

Percutaneous absorption

Parameter:

percentage

Absorption:

ca. 0.6 %

Remarks on result:

other:

Conversion factor human vs. animal skin:

See below

Any other information on results incl. tables

Key parameters:

- Prior to the determination of the percutaneous absorption of TMPTA, the permeation coefficient (Kp) for tritiated water was determined in human skin membranes. Skin membranes with a Kp value below the cut-off value of 2.5×10-3 cm.h-1 were selected for the study.

- The solubility of TMPTA in water was reported to be ca. 500 mg.L-1. In the study, the receptor fluid, PBS containing 0.01% sodium azide (w/v), was supplemented with 6% PEG (w/v), pH 7.2 to improve solubility of the test substance in the receptor fluid. The solubility of TMPTA in the receptor fluid was determined in this study and found to be ca. 93 μg.mL-1. The maximum absorption into the receptor fluid was 14.5 μg (i.e. 22.6 μg.cm-2, replicate A-3) in ca. 40 mL over 24 h, i.e. 0.36 μg.mL-1. Therefore, the solubility of TMPTA in the receptor fluid was considered sufficient. Furthermore, in the flow-through cells used, the volume of the receptor fluid in the receptor chamber beneath the skin was ca. 0.2 mL, which at a flow rate of ca. 1.8 mL.h-1, was replenished continuously (9 times per hour). Thus, it was assured that the rate of diffusion into the receptor fluid did not become a rate-limiting step (i.e. sink conditions were maintained).

- The mean absorption of TMPTA from the test group A into the receptor fluid over the 24 h study duration was 14.6 μg.cm-2, representing 0.16% of the applied dose.

- The mean maximal flux for the absorption of TMPTA through human skin was 0.77 μg.cm- 2.h-1 and the lag time was 0.2 h.

- The mean total absorption, defined as the compound-related radioactivity present in the receptor fluid, the receptor compartment wash and the skin membranes (excluding tape strips) was 0.32 ± 0.12% of the applied dose. The amount in tape strips 3 -15 was 0.28 +/- 17. The mean potentially absorbed dose, which is defined as the compound-related radioactivity present in the receptor fluid, the receptor compartment wash, the skin membranes and the stratum corneum (except for the first 2 tape strips) was 0.60 +/- 0.26 % of the applied dose.

- The mean recovery of TMPTA in human skin was 101.9 +/- 1.1%. For the dose formulation tested on human skin, less than 75 % of the absorption of TMPTA in the receptor fluid over 24 hours occurred within half of the study duration (i.e. 12 hours).

- For risk assessment, in agreement with the EFSA Scientific Opinion behind the revision of the Guidance Document on Dermal Absorption (2012), it is considered appropriate to include the tape strips (except the first 2 tape strips) in the calculations of the total absorption values (i.e. the potentially absorbed dose).

Applicant's summary and conclusion

Conclusions:

The percutaneus absorption of TMPTA through human skin membranes was evaluated in accordance to OECD 428. Using radioactive labelled TMPTA ([14C] TMPTA) for evaluation of penetration through human skin, the mean total absorption, defined as the compound-related radioactivity present in the receptor fluid, the receptor compartment wash and the skin membranes (excluding tape strips) was 0.32 ± 0.12% of the applied dose. The mean potentially absorbed dose, which is defined as the compound-related radioactivity present in the receptor fluid, the receptor compartment wash, the skin membranes and the stratum corneum (except for the first 2 tape strips) was 0.60 +/- 0.26 % of the applied dose.

Executive summary:

The percutaneus absorption of Trimethylolpropane triacrylate (TMPTA) through human skin membranes was evaluated in accordance to OECD 428. TMPTA was tested as neat compound, which represents the maximal concentration possible when handling the neat compound. The objective of the study was to elucidate the extent of percutaneous absorption of the compound-related radioactivity. The contact time was 8 hours, i.e. a normal working day and the post exposure time was 16 hours (sampling duration was 24h). In addition to the amount of radioactivity in the receptor fluid, the residues remaining in/on the skin membranes and in the stratum corneum (16 h post exposure) were determined. The study was performed in flow-through diffusion cells.

Using radioactive labelled TMPTA ([14C] TMPTA) for evaluation of penetration through human skin, the mean absorption of TMPTA into the receptor fluid over the 24 h study duration was 14.6 μg.cm-2, representing 0.16% of the applied dose. The mean maximal flux for the absorption of TMPTA through human skin was 0.77 μg.cm- 2.h-1 and the lag time was 0.2 h.

The mean total absorption, defined as the compound-related radioactivity present in the receptor fluid, the receptor compartment wash and the skin membranes (excluding tape strips) was 0.32 ± 0.12% of the applied dose. The mean potentially absorbed dose, which is defined as the compound-related radioactivity present in the receptor fluid, the receptor compartment wash, the skin membranes and the stratum corneum (except for the first 2 tape strips) was 0.60 +/- 0.26 % of the applied dose.