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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin irritation / corrosion
Remarks:
in vitro
Type of information:
experimental study
Adequacy of study:
key study
Study period:
November 3, 2014 - November 7, 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)
Version / remarks:
(2013)
Deviations:
no
Qualifier:
according to
Guideline:
other: EU Method B.40 BIS (In Vitro Skin Corrosion: Human Skin Model Test)(2008)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Strontium tartrate (anhydrous)
- Appearance: Off-white powder

Test animals

Details on test animals and environmental conditions:
All incubations, with the exception of the test substance incubation of 3 minutes at room temperature, were carried out in a controlled environment, in which optimal conditions were a humid atmosphere of 80 - 100% (actual range 80 - 88%), containing 5.0 ± 0.5% CO2 in air in the dark at 37.0 ± 1.0°C (actual range 36.6 - 36.7°C).

Test system

Vehicle:
unchanged (no vehicle)
Amount / concentration applied:
TEST MATERIAL
- Amount applied: 25 mg
- The test material was spread to match the size of the tissue

NEGATIVE CONTROL
- Amount applied: 50 µL Milli-Q water (Millipore Corp., Bedford, Mass., USA)

POSITIVE CONTROL
- Amount applied: 50 µL KOH (Merck, Darmstadt, Germany)
- Concentration (if solution): 8N
Duration of treatment / exposure:
- 3 minutes
- 1 hour
Details on study design:
TEST FOR REDUCTION OF MTT
The test substance was checked for possible direct MTT reduction before the study was started. To assess the ability of the test substance to reduce MTT, 122.5 mg of the test substance was added to 1.2 mL MTT medium. The mixture was incubated for approximately 1 hour at 37.0 ± 1.0°C. A negative control, sterile Milli-Q water was tested concurrently.

APPLICATION/TREATMENT
The test was performed on a total of 4 tissues per test substance together with a negative control and positive control. Two tissues were used for a 3-minute exposure to the test substance and two for a 1-hour exposure. The skin was moistened with 25 µL Milli-Q water (Millipore Corp., Bedford, Mass., USA) to ensure close contact of the test substance to the tissue and 25 mg of the solid test substance (glass weight boat) was added into the 6-well plates on top of the skin tissues. The remaining tissues were treated with 50 µL Milli-Q water (negative control) and with 50 µL 8N KOH (positive control), respectively.

REMOVAL OF TEST SUBSTANCE
- Washing: phosphate buffered saline (Invitrogen Corporation, Breda, The Netherlands)
- Time after start of exposure: 3 minutes and 1 hour
- Remaining test substance was removed with a cotton wool swab
- At the 1-hour exposure time one of the skin tissues was damaged during removal of the test substance with the cotton wool swab
- Rinsed tissues were kept in 24 well plates on 300 µL DMEM medium until 6 tissues (= one application time) were dosed and rinsed.

SCORING SYSTEM
- Percentage viability. Cell viability was calculated for each tissue as percentage of the mean of the negative control tissues. Skin corrosion potential of the test substance was classified according to remaining cell viability following exposure of the test substance with either of the two exposure times.
- The DMEM medium was replaced by 300 µL MTT-medium and tissues were incubated for 3 hours at 37°C in air containing 5% CO2. After incubation the tissues were washed with PBS and formazan was extracted with 2 mL isopropanol (MatTek corporation) over night at room temperature. The amount of extracted formazan was determined spectrophotometrically at 540 nm in triplicate with the TECAN Infinite® M200 Pro Plate Reader.

Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
other: Percentage viability
Basis:
other: Percentage of control
Time point:
other: 3 minutes
Score:
83
Remarks on result:
other: Negative control: 100%; Positive control: 11%
Irritation parameter:
other: Percentage viability
Basis:
other: Percentage of control
Time point:
other: 1 hour
Score:
61
Remarks on result:
other: Negative control: 100%; Positive control: 7%

Any other information on results incl. tables

- Mean absorption (duplicate exposures are indicated by A and B):

 

3-minute application

1-hour application

A (OD540)

B (OD540)

Mean

(OD540)

SD

A (OD540)

B (OD540)

Mean

(OD540)

SD

Negative control

1.642

1.620

1.631

±

0.015

1.533

1.515

1.524

±

0.013

Test substance

1.517

1.183

1.350

±

0.236

0.726

1.135

0.930

±

0.289

Positive control

0.198

0.146

0.172

±

0.036

0.112

0.098

0.105

±

0.010

SD = Standard deviation

In this table the values are corrected for background absorption (0.042). Isopropanol was used to measure the background absorption.

- Strontium tartrate (anhydrous) was checked for possible direct MTT reduction by adding the test substance to MTT medium. Because no colour change was observed it was concluded that Strontium tartrate (anhydrous) did not interact with MTT.

 

- Mean tissue viability:

 

3-minute application

viability (percentage of control)

1-hour application

viability (percentage of control)

Negative control

100

100

Strontium tartrate (anhydrous)

83

61

Positive control

11

7

 

Applicant's summary and conclusion

Interpretation of results:
other: not corrosive
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
In an in vitro skin corrosion test using a human three dimensional epidermal model, conducted according to OECD 431 and GLP principles, it was determined that the substance is not corrosive to skin.
Executive summary:

In an in vitro skin corrosion test using a human three dimensional epidermal model (EpiDerm (EPI-200)), conducted according to OECD 431 and GLP principles, the possible corrosive potential of the substance was tested through topical application for 3 minutes and 1 hour. Reliable negative and positive controls were included. Skin tissue was moistened with 25 µL of Milli-Q water and 25 mg of the substance was applied directly on top of the skin tissue. Skin corrosion is expressed as the remaining cell viability after exposure to the test substance. The relative mean tissue viability obtained after 3-minute and 1-hour treatments with the substance compared to the negative control tissues was 83% and 61%, respectively. Because the mean relative tissue viability was not below 50% after the 3-minute treatment and not below 15% after the 1-hour treatment, the substance is considered to be not corrosive to skin.