Reaction products of imine and acrylate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation / corrosion

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: OECD guideline and EU method. GLP study.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 430 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test Method (TER))

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Reaction products of imine and acrylate

Molecular formula:

Not applicable. UVCB substance

IUPAC Name:

Reaction products of imine and acrylate

Details on test material:

- Physical state: Clear amber liquid.
- Lot/batch No.: 3017/20461
- Storage condition of test material: in a cool, airy places away from strong alkali, acids, ignition sources, direct sunlight.

Test animals

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Center for Experimental Medicine in Medical University of Silesia in Katowice.
- Age at study initiation: 28-day old
- Housing: Animals were kept in cage with dimensions (length x width x height): 58 x 37 x 21 cm; with plastic bottom and wired lid.
- Diet (e.g. ad libitum): standard granulated "Murigran" fodder, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: 3 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 22 ºC
- Humidity (%): 55 – 80 %
- Air changes (per hr): about 16 times/hour
- Photoperiod (hrs dark / hrs light): 12 hours light / 12 hours dark

Test system

Type of coverage:

other: in vitro study

Preparation of test site:

clipped

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): The test item was applied evenly to the disc, which was placed inside the tube in volume of 150 μl.

Duration of treatment / exposure:

Test item and control substances were applied for 2 hours (36% hydrochloric acid) or 24 hours (distilled water and test item) at 21-22°C. Then the test item and control substances were removed by washing off with a jet of tap water at temperature up to 30°C.

Number of animals:

Two animals

Details on study design:

PREPARATION OF THE SKIN DISCS
The animals were subjected to euthanasia by intraperitoneal administration of morbital in amount of 200 mg/kg b.w. before taking skin discs.
After euthanasia the dorso-lateral skin of each animal was then removed and stripped of excess subcutaneous fat by carefully peeling it away from the skin by using paper towel, and the skin discs were cut out by scalpel. 11 skin discs (2 for quality control procedure, 9 for main testing) with a diameter 20-mm each, were obtained from a single rat skin.

NUMBER OF REPLICATES
Three skin discs, which were obtained from skin of each animal, were used for the test item and control substance (three repetitions).

CONTROLS
The positive (36% hydrochloric acid) and negative (distilled water) controls were used concurrently in each study to ensure adequate performance of the experimental model.

Results and discussion

Any other information on results incl. tables

RESULTS OF ELECTRICAL RESISTANCE (TER) MEASUREMENT

As a quality control procedure, before start of the experiment, the electrical resistance of two skin discs was measured for each animal skin (animal No 1 and No 2). In each case skin discs had resistance values greater than 10 kΩ, so the other skin discs of animals could be used in the experiment.

The mean TER results can be accepted because concurrent positive and negative control values felt within the ranges acceptable for the method (the mean TER results for 36% hydrochloric acid amounted to: 0.83 and 0.75 kΩ; for distilled water amounted to: 20.24 and 21.29 kΩ).

The mean TER results of the skin discs treated with the test item were equal to: 39.61 kΩ (Animal No 1) and 47.49 kΩ (Animal No 2).

The individual values of TER measurements were very diversified and they ranged from 28.47 to 71.23 kΩ which resulted from the deposition of the test item on the treated surface of the skin discs. Directly after termination of 24-hour exposure period and attempt to remove the test item by washing it off with a stream of tap water, the TER values ranged from 103.12 to 132.47 kΩ.

GROSS EVALUATION OF THE TREATED SKIN DISCS

During gross examination of the skin discs from positive control, skin perforation in all cases was stated, whereas in negative control and on the skin discs treated with the test item, no changes on the skin surface were stated. A hard, close-fitting, transparent layer of test item was formed on the surface of each treated skin disc.

RESULTS OF SULFORHODAMINE B CONTENT IN THE DISCS

Concurrent mean disc dye content values of positive and negative control were equal to: for 36% HCl –64.29 μg/disc (Animal No 1) and 70.32 μg/disc (Animal No 2), for distilled water 27.75 μg/disc (Animal No 1) and 29.35 μg/disc (Animal No 2). These values felt within the ranges acceptable for the method.

The mean disc dye content results of the skin discs treated with the test item were equal to 158.49 μg/disc (Animal No 1) and 156.28 μg/disc (Animal No 2). The high content of the dye in the treated skin disc may be result of Sulforhodamine B binding by the layer of the test item covering the discs.

HISTOPATHOLOGICAL EVALUATION OF THE TREATED DISCS

During histopathological examination of the treated discs no pathological changes were stated. Histopathological examination did not confirm the corrosion of test item.

Applicant's summary and conclusion

Interpretation of results:

other: not corrosive

Remarks:

Criteria used for interpretation of results: EU

Conclusions:

On the ground of the study, the test item does not comply with the criteria of the method and therefore based on the results we can not clearly conclude that the test item is corrosive.

Executive summary:

In vitro skin corrosion: Transcutaneous electrical resistance test (TER) was performed in order to obtain information on health hazard due to superficial influence of the test substance on skin. Skin discs which were used for the experiment, were obtained from 28-day-old animals. As a quality control procedure, before start of the experiment, the electrical resistance of two skin discs was measured from each animal skin. In each case the skin discs showed resistance values greater than 10 kΩ, so the other of the skin discs of animals could be used in the experiment. The undiluted test item was applied uniformly to the epidermal surface of skin disc placed inside a tube. The positive control (36% hydrochloric acid) and negative control (distilled water) were used concurrently in the study. Three skin discs of each animal were used for the test item and three for control materials. Test item and control substances were applied for 2 hours (36% hydrochloric acid) or 24 hours (distilled water and test item) at 21-22°C, and then removed by washing off with a jet of tap water. During 24-hour exposure period the test item upon skin contact took the form of a solid. A hard, close-fitting, transparent layer of test item was formed on the surface of each treated skin disc. In order to remove the test item the discs were soaked in tap water (at about 30 ° C) for 1 hour and shaken every few minutes. Prior to measurement of the electrical resistance, the surface tension of the skin was reduced by adding a sufficient volume of 70% ethanol to cover the epidermis. After few seconds, the ethanol was removed from the tube and the tissue was then hydrated by the addition of 3 mL MgSO4 solution (154 mM). Low-voltage, alternating current LCR Databridge Model 6401 bridge was used to measure the electrical resistance of the skin in kΩ, by placing databridge electrodes on either side of the skin disc. After transcutaneous electrical resistance test (TER), MgSO4 solution was removed from the tube, and the

skin discs were then subjected to gross evaluation in order to reveal possible damage. The dye binding procedure was necessary in this case, since the all TER values for the test item were very diversed.

On the ground of the study, the test item does not comply with the criteria of the method and therefore based on the results we can not clearly conclude that the test item is corrosive.