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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin corrosion: in vitro / ex vivo
Remarks:
in vitro
Type of information:
experimental study
Adequacy of study:
key study
Study period:
April 25, 2016- April 30, 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report Date:
2016

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 430 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test Method (TER))
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: crystalline

In vitro test system

Test system:
isolated skin discs
Source species:
rat
Cell type:
other: dorso lateral skin discs stripped of excess subcutaneous fat
Cell source:
other: skin obtained from two female donors
Source strain:
Wistar
Details on animal used as source of test system:
SOURCE ANIMAL
- Source: skin discs were collected from 2 female rats (outbred) obtained from the conventional husbandry of laboratory animals of the Centre for Experimental Medicine, Medical University in Katowice (reg. No. 0053)
- Sex: female
- Age at study initiation (in days): 21 days old
- Weight at study initiation: not specified
- Housing: both rats were kept in a plastic cage covered with a wire bar lid. Conditions: air temperature: 20-21ºC; relative air humidity: 28-51%; lighting cycle: 12 hours light/12 hours dark; air exhange: about 13 times/hour.
- Diet : ad libitum access to the “Murigran” (batch number 1/16) standard granulated laboratory fodder produced by Wytwórnia Koncentratów i Mieszanek Paszowych AGROPOL, Motycz
- Water: ad libitum drinking water (tap water)
- Acclimation period: 4 days
Justification for test system used:
OECD Guideline 430 is based on the rat skin transcutaneous electrical resistance (TER) test method, which utilizes skin discs to identify corrosives by their ability to produce a loss of normal stratum corneum integrity and barrier function.
Vehicle:
unchanged (no vehicle)
Details on test system:
SKIN DISC PREPARATION
- Procedure used: dorso-lateral skin of each animal (two female Wistar rats of 29 days old) was removed and stripped of excess subcutaneous fat by carefully peeling it away from the skin using a paper towel. Each skin disc was placed over one of the ends of a PTFE (polytetrafluoroethylene) tube, ensuring that the epidermal surface was in contact with the tube. A rubber ‘O’ ring was press-fitted over the end of the tube to hold the skin in place and excess tissue was trimmed away. The rubber ‘O’ ring was then carefully sealed to the end of the PTFE tube with petroleum jelly. The tube was supported by a spring clip inside a receptor chamber containing MgSO4 solution (154 mM). The skin disc should be fully submerged in the MgSO4 solution. As many as 11 skin discs with a diameter of 20-mm each were obtained from a single rat skin. Two of them were used to control the quality of the procedure, whereas the remaining nine were used for the purpose of the experiment.
- Quality control for skin discs: Electrical resistance obtained with two of the isolated skin discs should be greater than 10 kΩ

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 21-22ºC

REMOVAL OF TEST MATERIAL AND CONTROLS
- Number of washing steps: 1 (the test item and the control items were removed by washing with a jet of tap water at up to 30°C)
- Observable damage in the tissue due to washing: no.
- Modifications to validated SOP: no.

NUMBER OF INDEPENDENT TESTING RUNS / EXPERIMENTS TO DERIVE FINAL PREDICTION: 2

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be corrosive to skin if the mean TER value obtained for the test item is less than or equal to 5 kΩ and the skin disk is obviously damaged.
- The test substance is considered to be non-corrosive to skin if the mean TER value obtained for the test substance is greater than 5 kΩ or if the mean TER value is less than or equal to 5 kΩ and the skin disc is showing no obvious damage.
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): the test item (ground to a powder) was moistened with 150 µl distilled water and uniformly applied in a sufficient amount to ensure that the whole surface of the epidermis was covered.

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 150 µl of distilled water

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 150 µl of 10M hydrochloric acid
Duration of treatment / exposure:
24 hours
Duration of post-treatment incubation (if applicable):
not apllicable
Number of replicates:
three replicates obtained for each rat (two female) were used for the test item and for each control item.

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
transcutaneous electrical resistance (in kΩ)
Run / experiment:
#1 (mean value)
Value:
18.26
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Remarks:
14.97 kΩ
Positive controls validity:
valid
Remarks:
0.91 kΩ
Remarks on result:
no indication of irritation
Irritation / corrosion parameter:
transcutaneous electrical resistance (in kΩ)
Run / experiment:
# 2 (mean value)
Value:
15.12
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Remarks:
16.89 kΩ
Positive controls validity:
valid
Remarks:
0.87 kΩ
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
OTHER EFFECTS:
- Visible damage on test system: the gross examination showed no changes on the surface of the treated skin discs.

DEMONSTRATION OF TECHNICAL PROFICIENCY:
- Yes (study conducted from May to November 2014).

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes. TER values within the acceptable range, no changes on surface of the treated skin discs.
- Acceptance criteria met for positive control: yes. TER values within acceptable range and skin discs were perforated.
- Acceptance criteria met for variability between replicate measurements: yes, the difference of TER means were below 5 ± 0,5 kΩ.

Any other information on results incl. tables

Table 1. Results of the control transcutaneous electrical resistance test (TER): skin discs accepted if TER value > 10  (kΩ)

Animal number

Skin disc number

TER value (kΩ)

1

1

14.31

2

13.03

2

1

15.01

2

14.17

 

Table 2. Results of the transcutaneous electrical resistance test (TER)

Animal

number

Tested substance

Skin disc number

TER value (kΩ)

Mean TER value

± SD (kΩ)

Control ranges

(kΩ)

1

Positive control –

10M HCl

1

0.91

0.91± 0.01

       0.5 -1.0

2

0.92

3

0.90

Negative control –

Distilled water

1

14.78

14.97 ± 0.20

       10 -25

2

14.95

3

15.17

Test item

1

18.23

18.26 ± 0.32

       

2

17.96

3

18.59

2

Positive control –

10M HCl

1

0.88

0.87± 0.01

       0.5 -1.0

2

0.87

3

0.86

Negative control –

Distilled water

1

17.35

16.89 ± 0.52

       10 -25

2

17.01

3

16.32

Test item

1

15.36

15.12 ± 0.21

       

2

14.98

3

15.01

 

Table 3. Gross changes on the surface of the treated skin discs.

Animal

number

Tested substance

Skin disc number

Gross changes

1

Positive control –

10M HCl

1

Perforation

2

Perforation

3

Perforation

Negative control –

Distilled water

1

No changes

2

No changes

3

No changes

Test item

1

No changes

2

No changes

3

No changes

2

Positive control –

10M HCl

1

Perforation

2

Perforation

3

Perforation

Negative control –

Distilled water

1

No changes

2

No changes

3

No changes

Test item

1

No changes

2

No changes

3

No changes

Applicant's summary and conclusion

Interpretation of results:
other: non-corrosive
Remarks:
EU criteria
Conclusions:
The mean TER values for the test item were higher than 5 kΩ and there were not any visible changes on the skin discs. Therefore, the test item is non-corrosive to skin.
Executive summary:

A transcutaneous electrical resistance test (TER) was performed on the test item according to OECD TG 430 (GLP study). Skin discs used in the experiment were obtained from two 29-day-old female Wistar rats. The test item (ground to a powder) was uniformly applied to the epidermal surface of these skin discs placed inside a tube. Positive (10M hydrochloric acid) and negative (distilled water) controls were conducted concurrently. Three skin discs obtained from each animal were used for the test item and three for each control item. The skin discs were incubated for 24 hours and kept at 21-22°C. Then, substances were removed by washing with a jet of tap water. Prior to measuring the electrical resistance, the surface tension of the skin discs was reduced by adding a sufficient volume of 70% ethanol. After removing the ethanol the tissue was hydrated by the addition of 3 ml of a solution of MgSO4 (154 mM). A LCR 6401 low-voltage, alternating current databridge was used to measure the electrical resistance of the skin discs (in kΩ) by placing the databridge electrodes on either side of the skin disc. After the measurements, the skin discs were subjected to a gross examination in order to reveal possible damage. The mean TER results for the skin discs treated with the test item were 18.26 kΩ (animal no. 1) and 15.12 kΩ (animal no. 2), higher than the 5 kΩ cut-off value. Positive and negative control values felt within the acceptable ranges for the method. Gross examinations of the skin discs treated with the test item did not reveal any changes. On the basis of these conditions, the test item was considered to be non-corrosive to skin.