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EC number: 204-445-3 | CAS number: 121-03-9
Skin irritation / corrosion
Administrative data
- Endpoint:
- skin irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 010
- Report date:
- 2010
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)
- GLP compliance:
- yes
Test material
- Reference substance name:
- 4-nitrotoluene-2-sulphonic acid
- EC Number:
- 204-445-3
- EC Name:
- 4-nitrotoluene-2-sulphonic acid
- Cas Number:
- 121-03-9
- Molecular formula:
- C7H7NO5S
- IUPAC Name:
- 2-methyl-5-nitrobenzenesulfonic acid
Constituent 1
In vitro test system
- Test system:
- human skin model
- Details on animal used as source of test system:
- EpiDerm-Test
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- The EpiDermTM model consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multi layered, highly differentiated model of the human epidermis. It consists of organized basal, spinous and granular layers, and a multi-layered stratum corneum containing intercellular lamellar lipid layers arranged in patterns analogous to those found in vivo. The EpiDermTM tissues (surface 0.6 cm²) are cultured on specially prepared cell culture inserts (MILLICELLs®, 10 mm Ø) and commercially available as kits (EpiDerm™ 200), containing 24 tissues on shipping agarose.
TEST PROCEDURE
- For the corrosion test two tissue samples were uses for 3 minutes and 1 hour, the irritaion test was perfomed with tree tissue sampels, which were incubated with the test substance for 1 hour followed by a 42-hours post-incubation period.
- After 3 minutes (room temperature) or after 1 hour (incubator) tissue contact with the test material, the tissues were washed with buffer to remove residual test material. Rinsed tissues were kept in 24-well plates (holding plates) at room temperature on assay medium until all tissues per application time were dosed and rinsed. The assay medium was then replaced by MTT solution and tissues were incubated for 3 hours. After incubation tissues were washed with buffer and formazan was extracted with Isopropanol over night at room temperature. The optical density at a wavelength of 570 nm (OD570) of extracted formazan was determined spectrophotometrically and cell viability was calculated for each tissue as % of the mean of the negative control tissues. Skin corrosivity potential of the test material was classified according to remaining cell viability obtained after test material treatment with either of the 2 exposure times.
CONTROLS
- Negative control (NC): Doubly distilled water
- Positive control (PC): 8 N potassium hydroxyde (Sigma-Aldrich, Munich, Germany)
ASSAY ACCEPTANCE CRITERIA:
- Negative control (NC ): mean OD570 of the NC per exposure time 2.5
- Positive control (PC ): mean relative tissue viability of the 3 min positive control is <= 30 %
- Tissue variability: The inter tissue variability is considered to be acceptable if the difference of the OD570 values of the two tissues is <= 0.3 - Amount/concentration applied:
- - Amount(s) applied (volume or weight with unit): 25 µl of the solid test substance
- Duration of treatment / exposure:
- 3 minutes and 1 hour
Results and discussion
In vitro
Resultsopen allclose all
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 3 min / corrosion test
- Value:
- 101
- Vehicle controls validity:
- not examined
- Negative controls validity:
- not specified
- Positive controls validity:
- not specified
- Remarks on result:
- no indication of irritation
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- irritation test
- Value:
- 7
- Vehicle controls validity:
- not examined
- Negative controls validity:
- not specified
- Positive controls validity:
- not specified
- Remarks on result:
- positive indication of irritation
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 1 hour/corrosion test
- Value:
- 17
- Vehicle controls validity:
- not examined
- Negative controls validity:
- not specified
- Positive controls validity:
- not specified
- Remarks on result:
- positive indication of irritation
- Other effects / acceptance of results:
- Based on the observed results and applying the evaluation criteria cited, it was concluded, that 4-Nitrotoluene-2-sulphonic acid shows a skin irritation potential in the EpiDerm TM skin corrosion/irritation test under the test conditions chosen
Any other information on results incl. tables
Corrosion test
|
|
Exposure: 3 min |
Exposure: 1 hour |
||||
Test Substance |
|
Tissue 1 |
Tissue 2 |
Mean |
Tissue 1 |
Tissue 2 |
mean |
NC |
Mean OD570 |
1.6233 |
1.8758 |
1.7496 |
1.5827 |
1.8112 |
1.6969 |
|
Viability [% of NC] |
92.8 |
107.2 |
100 |
93.3 |
106.7 |
100 |
09/0641-1 |
Mean OD570 |
1.9028 |
1.6308 |
1.7668 |
0.2857 |
0.2772 |
0.2814 |
|
Viability [% of NC] |
108.8 |
93.2 |
101 |
16.8 |
16.3 |
17 |
PC |
Mean OD570 |
0.3493 |
0.3223 |
0.3358 |
0.1902 |
0.1907 |
0.1904 |
|
Viability [% of NC] |
20.0 |
18.4 |
19 |
11.2 |
11.2 |
11 |
Irritation Test
Test Substance |
|
Tissue 1 |
Tissue 2 |
Tissue 3 |
Mean |
SD |
NC |
Mean OD570 |
1.8535 |
2.3000 |
* |
2.0768 |
|
|
Viability [% of NC] |
89.3 |
110.7 |
* |
100 |
15.20 |
09/0641-1 |
Mean OD570 |
0.1445 |
0.1385 |
0.1635 |
0.1488 |
|
|
Viability [% of NC] |
7.0 |
6.7 |
7.9 |
7 |
0.63 |
PC |
Mean OD570 |
0.1005 |
0.1140 |
0.1000 |
0.1048 |
|
|
Viability [% of NC] |
4.8 |
5.5 |
4.8 |
5 |
0.38 |
* For the NC two tissues were valid for evaluation, only (details are available with the raw data)
Applicant's summary and conclusion
- Interpretation of results:
- other: classified as a skin irritant according to the CLP Regulation EC No. 1272/2008
- Conclusions:
- The substance is a skin irritant (Cat.2).
- Executive summary:
The skin irritation potential of the test substance was evaluated following the OECD 431.
For the corrosion test the samples were incubated with the test substance for 3 minutes and 1 hour, for the irritation test the samples were incubated with the test substance for 1 hour followed by a 42 -hours post-incubation period. Tissue destruction was determined by measuring the metabolic acitvity of the tissue after exposure/post-incubation using a colorimetric test. The reduction of mitochondrial dehydrogenase activity, measured by reducted formazan production after incubation with a tetrazolium salt (MTT) was chosen as suitable endpoint.The mean viability of the test substance treated tissues determined after an exposure period of 3 minutes was 101%, and it was 17% after an exposure period of 1 hour. The mean viability of the test-substnce treated tissues determined after an exposure period of 1 hour with about 42 hours post-incubation was 7%.
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