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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Remarks:
other: in vitro
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2012-02-13 to 2012-02-17
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
other: EU Method b.46 (Skin Irritation)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany

Test material

Constituent 1
Chemical structure
Reference substance name:
Sodium 4-(methoxycarbonyl)phenolate
EC Number:
225-714-1
EC Name:
Sodium 4-(methoxycarbonyl)phenolate
Cas Number:
5026-62-0
Molecular formula:
C8H8O3.Na
IUPAC Name:
methyl 4-hydroxybenzoate
Details on test material:
Name: Sodium methyl-4-hydroxybenzoate
CAS No.: 5026-62-0
Chemical Name: sodium 4-(methoxycarbonyl)phenolate
Physical State/Colour: solid/white
Storage Conditions: room temperature

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
other: EpiDerm( MatTek) rganotypic reconstructed three-dimensional model of the human epidermis
Cell source:
other: EpiDerm( MatTek)
Details on animal used as source of test system:
organotypic reconstructed three-dimensional model of the human epidermis
Justification for test system used:
Ths test method is able to detect chemicals that cause skin irritation, i.e. produce reversible daamage to the skin and allows for hazard identification in accordance with UN GHS category 2.
Vehicle:
other: Dulbecco's Phosphate Buffrered Saline (DPBS)
Details on test system:
This test used the EpiDerm TM reconstructed human epidermis model (MatTk) which consists of human keratinocytes (NHEK) and therefore represents in vitro the target organ of the species of ntrest and closely mimics the biochemical and physiological properties of the upper parts of the human, i.e. the epidermis.
Control samples:
yes, concurrent negative control
yes, concurrent vehicle
yes, concurrent positive control
Amount/concentration applied:
25 mg test substance in 25 µL DPBS
Duration of treatment / exposure:
60 ± 1 min
Duration of post-treatment incubation (if applicable):
Post-treatment incubation period: 42 ± 4 h
Number of replicates:
The test was performed in triplicates for each treatment and control group.

Test system

Details on study design:
3 replicate tissues are dosed with the test item, the negative control (DPBS) and the positive control (5% SDS), respectively. The tissues are incubated for 35 ± 1 minutes in a humidified incubator (37 ± 1 °C, 5 ± 1% CO2, 95% RH) and further 25 minutes at room temperature. Then the test item and the controls are rinsed off with DPBS and the tissues are post-incubated for 24 ± 2 h and, after medium renewal, for further 18 ± 2 h. Then the tissues are stained via MTT for 3 hours. Isopropanol extracts are measured photometrically at 550 nm.

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
Negative control / 60 min exposure / mean
Value:
>= 100
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
Positive control / 60 min exposure / mean
Value:
ca. 2.2
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks:
5 % SDS
Remarks on result:
positive indication of irritation
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
test item/ 60 min exposure/ mean
Value:
ca. 6.2
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
positive indication of irritation
Other effects / acceptance of results:
The test item showed no direct MTT reducing capability and no colouring potential.

Any other information on results incl. tables

Name

Negative Control

Positive Control

Test Item

Tissue

1

2

3

1

2

3

1

2

3

corrected OD550

2.171

2.167

2.186

0.052

0.048

0.044

0.094

0.172

0.132

2.111

2.203

2.114

0.050

0.046

0.048

0.099

0.176

0.134

mean corrected OD550
of the duplicates

2.141

2.185

2.150

0.051

0.047

0.046

0.097

0.174

0.133

mean corrected OD550
of 3 replicate tissues

2.159*

0.048

0.134

mean SD OD550

0.038

0.003

0.035

tissue viability [%]

99.2

101.2

99.6

2.4

2.2

2.1

4.5

8.1

6.2

SD tissue viability [%]***

1.1

0.1

1.8

CV [% viability]

1.1

5.5

28.7

mean tissue viability [%]

100.0

2.2**

6.2

*       Corrected mean OD550 of the negative control corresponds to 100% absolute tissue viability.

**      mean relative tissue viability of the three positive control tissues is  20%

***    standard deviation (SD) obtained from the three concurrently tested tissues is < 18%

Applicant's summary and conclusion

Interpretation of results:
irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
CLP: Skin Irrit. 2, H315
DSD: Xi, R38
Executive summary:

In the present study the skin irritant potential of Sodium methyl-4-hydroxybenzoate was analysed. The EpiDerm TM-Standard Model (EPI-200 TM), a reconsituted three-dimensional human epidermis model, was used as a replacement of the Draize skin irritation test (OECD 404) to distinguish between UN GHS and EU CLP "Category 2" skin irritating test substances and not categorized test substances ("no category") which may be considered as non-irritant. Hereby, the test item was applied topically, cytotoxicity is expressed as the reduction of mitochondrila dehydrogenase activity measured by formazan production from MTTafter a 60 min exposure and 42 h post incubation period and compared to those of the concurrent negative controls. In this study under the given conditions the test item showed irritant effects. The relative mean tissue viability after 60 min of exposure and 42 h post incubation was < 50%. The test item is therefore classified as "irritant" in accordance with UN GHS and EU CLP "Category 2".

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