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Description of key information

The test substance was found to be corrosive to skin, therefore, no test for eye irritation/corrosion has to be conducted.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
from 2015-06-12 to 2015-07-06
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
GLP and OECD Guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Version / remarks:
2013
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test system:
human skin model
Remarks:
Reconstructed Human Epidermis model
Details on animal used as source of test system:
Test system:
Epi-200- SIT Kit (Lot No.: 21678)
MTT-100 Assay Kit

Epi-200 SIT kits and MTT-100 assay diluents were purchased from MatTek Corporation (82105 Bratislava, Slovakia). The EpiDermTM tissue 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 EpiDerm™ tissues (surface 0.6 cm²) are cultured on specially prepared cell culture inserts (MILLICELLs, d=10 mm).
EpiDerTM tissues were shipped with cool packs on medium-supplemented agarose gels in a 24-well plate and reached Envigo CRS on June 30, 2015. On day of receipt the pre-incubation phase of the tissues started.
Vehicle:
unchanged (no vehicle)
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
- Amount applied: 30 μL
Duration of treatment / exposure:
60 minutes
Number of replicates:
Three individual tissues were used for the test item and the positive control, respectively.
Observation period:
The colour of the test item/water mixture was observed during the whole incubation period (60 min). The measurement of the OD of the test item in water at 570 nm was not required and consequently not performed.
Details on study design:
SCORING SYSTEM:
The mean OD of the three negative control tissues was calculated after blank correction. This value corresponds to 100 % tissue viability in the current test. For each individual tissue treated with the test item or the positive control the individual relative tissue viability is calculated according to the following formula:
Relative viability [%] = (mean OD (test item/positive control) /mean OD (negative control)) * 100
For the test item and the positive control the mean relative viability ± rel. standard deviation of the three individual tissues was calculated and used for classification according to the following prediction model:
For the current test, an irritation potential of a test item according to EU classification H315 (according to regulation (EC) 1272/2008), and GHS category 2 according to UN GHS (published 2003, last (6th) revision 2015) is recommended, if the mean relative tissue viability of three individual tissues is reduced ≤ 50 % of the negative control.
Irritation / corrosion parameter:
other: relative absorbance ( % of negative control)
Run / experiment:
60 min
Value:
5.1
Remarks on result:
other: Basis: test item, mean of 3 tissues.
Irritation / corrosion parameter:
other: relative absorbance ( % of negative control)
Run / experiment:
60 min
Value:
6.5
Remarks on result:
other: Basis: positive control, mean of 3 tissues.
Other effects / acceptance of results:
The mean relative absorbance after treatment with the test item was 5.1 (% of negative control). For the positive control the mean relative absorbance was found to be 6.5 (% of negative control).
Interpretation of results:
Category 1B (corrosive) based on GHS criteria
Conclusions:
The test item was found to be irritating to skin according to UN GHS and EU CLP regulation.
Executive summary:

An in vitro study was performed to assess the irritation potential of the test item by means of the Human Skin Model Test. The test item did not reduce MTT (test for direct MTT reduction), and it did not change colour when mixed with deionised water (test for colour interference). Also its intrinsic colour was not intensive. Consequently, additional tests with freeze-killed or viable tissues were not necessary.

Each three tissues of the human skin model EpiDermTM were treated with the test item, the negative or the positive control for 60 minutes.

30 μL of the test item were applied to each tissue, and spread to match the surface of the tissue. 30 μL of either the negative control (DPBS) or the positive control (5 % SLS) were applied to each tissue.

After treatment with the test item the mean relative absorbance value decreased to 5.1 % compared to the relative absorbance value of the negative control. This value is below the threshold for irritancy of ≤ 50 %. Therefore, the test item is considered to possess an irritant potential.

In conclusion, it can be stated that in this study and under the experimental conditions reported, the substance is irritating to skin.

Endpoint:
skin corrosion: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
from 2015-09-03 to 2015-09-24
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
GLP and OECD Guidelien conform study
Qualifier:
according to guideline
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test system:
human skin model
Remarks:
EpiDerm TM Kit (human-derived epidermal keratinocytes )
Source species:
human
Cell type:
non-transformed keratinocytes
Details on animal used as source of test system:
Epi-200 kits and MTT-100 assays were purchased from MatTek Corporation (Bratislava, Slovakia). The EpiDerm™ tissue consisted of normal, human-derived epidermal keratinocytes which have been cultured to form a multilayered, highly differentiated model of the human epidermis. It consisted of organised 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 EpiDerm™ tissues (surface 0.63 cm²) are cultured on specially prepared cell culture inserts.
EpiDerm™ tissues were shipped at 4 °C on medium-supplemented agarose gels in a 24-well plate and reached Envigo CRS on 22 September 2015. On day of receipt the pre-incubation phase of the EpiDerm™ tissues started.
Vehicle:
unchanged (no vehicle)
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
- Amount applied: 50 µL
Duration of treatment / exposure:
3 min and 60 min
Number of replicates:
Duplicate tissues were treated with the test item, positive control or negative control.
Details on study design:
SCORING SYSTEM:
The mean OD value obtained for the duplicate tissues per test item was used to calculate a percent viability relative to the negative control, which is arbitrarily set at 100 %.
Irritation / corrosion parameter:
other: relative absorbance (% of negative control)
Run / experiment:
3 min
Value:
19.2
Remarks on result:
other: Basis: positive control
Irritation / corrosion parameter:
other: relative absorbance (% of negative control)
Run / experiment:
3 min
Value:
104.5
Remarks on result:
other: Basis: test item
Irritation / corrosion parameter:
other: relative absorbance (% of negative control)
Run / experiment:
60 min
Value:
5.2
Remarks on result:
other: Basis: positive control
Irritation / corrosion parameter:
other: relative absorbance (% of negative control)
Run / experiment:
60 min
Value:
8.6
Remarks on result:
other: Basis: tets item
Other effects / acceptance of results:
The test item is considered to be corrosive to skin:
The viability after 3 minutes exposure is greater than 50 % and the viability after 1 hour exposure is lower than than 15 %.
Interpretation of results:
Category 1B (corrosive) based on GHS criteria
Conclusions:
The test item is corrosive to skin according to EU CLP and UN GHS.
Executive summary:

An in vitro study was performed to assess the corrosive potential of the test substance by means of the Human Skin Model Test with EpiDerm tissue models. The test item did not reduce MTT (test for direct MTT reduction), and it did not change colour when mixed with deionised water (test for colour interference). Independent duplicate tissues of EpiDerm were exposed to either the test item, the negative control (deionised water) or the positive control (8.0 N KOH) for 3 minutes and 1 hour, respectively. 50 µL of the test item were dispensed directly onto duplicate EpiDerm tissue surface, and spread to match the surface of the tissue. Exposure to the positive control induced a decrease in the relative absorbance as compared to the negative control, both for the 3 minutes exposure period and for the 1 hour exposure period. The 1 hour exposure caused a decrease of the cell viability < 15 % of the negative control. The CV in the range 20 – 100 % viability between the tissue replicates is ≤ 30 %, thus the validity of the test system and the specific batch of the tissue models is confirmed. After exposure of the tissues to the test item the relative absorbance value did not decrease (104.5 %) after 3 minutes exposure. After 1 hour exposure the relative absorbance value was reduced to 8.6 %. The value for the 3 minutes exposure did not exceed the threshold for corrosivity, which is defined to be 50 %, but the value of 8.6 % felt below the threshold of 15 % for the 1 hour exposure. Therefore, the test item was considered to be corrosive.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (corrosive)

Eye irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Several study reports and publications are available assessing the skin irritating property of the test substance.

An in vitro study was performed to assess the irritation potential of the test item by means of the Human Skin Model Test. The test item did not reduce MTT (test for direct MTT reduction), and it did not change colour when mixed with deionised water (test for colour interference). Also its intrinsic colour was not intensive. Consequently, additional tests with freeze-killed or viable tissues were not necessary.

Each three tissues of the human skin model EpiDermTM were treated with the test item, the negative or the positive control for 60 minutes.

30 μL of the test item were applied to each tissue, and spread to match the surface of the tissue. 30 μL of either the negative control (DPBS) or the positive control (5 % SLS) were applied to each tissue.

After treatment with the test item the mean relative absorbance value decreased to 5.1 % compared to the relative absorbance value of the negative control. This value is below the threshold for irritancy of ≤ 50 %. Therefore, the test item is considered to possess an irritant potential.

In conclusion, it can be stated that in this study and under the experimental conditions reported, the substance is irritating to skin.

An in vitro study was performed to assess the corrosive potential of the test substance by means of the Human Skin Model Test with EpiDerm tissue models. The test item did not reduce MTT (test for direct MTT reduction), and it did not change colour when mixed with deionised water (test for colour interference). Independent duplicate tissues of EpiDerm were exposed to either the test item, the negative control (deionised water) or the positive control (8.0 N KOH) for 3 minutes and 1 hour, respectively. 50 µL of the test item were dispensed directly onto duplicate EpiDerm tissue surface, and spread to match the surface of the tissue. Exposure to the positive control induced a decrease in the relative absorbance as compared to the negative control, both for the 3 minutes exposure period and for the 1 hour exposure period. The 1 hour exposure caused a decrease of the cell viability < 15 % of the negative control. The CV in the range 20 – 100 % viability between the tissue replicates is ≤ 30 %, thus the validity of the test system and the specific batch of the tissue models is confirmed. After exposure of the tissues to the test item the relative absorbance value did not decrease (104.5 %) after 3 minutes exposure. After 1 hour exposure the relative absorbance value was reduced to 8.6 %. The value for the 3 minutes exposure did not exceed the threshold for corrosivity, which is defined to be 50 %, but the value of 8.6 % felt below the threshold of 15 % for the 1 hour exposure. Therefore, the test item was considered to be corrosive.

In a publication (Takenaka et al. 1968) the testing of several chemicals on the skin of human volunteers is described. The test substance was exposed to the skin sites under occlusion for 24 - 48 hours at a concentration of 0.05 - 0.5 % to 82 humans. As a result, no skin reactions were seen (refer to IUCLID6 Section 7.10.5).

 

Based on a weight of evidence approach, the substance is considered to be corrosive to skin at a concentration of 100 %. Therefore, in accordance with column 2 of Annex VII of the REACH Regulation, no study for eye irritation/corrosion has to be conducted.

Justification for classification or non-classification

Based on the above mentioned study results, the substance is considered to be classified for skin corrosion into the Category 1B or 1C under Regulation (EC) No 1272/2008 as amended for the tenth time in Regulation (EC) No 2017/776.