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Description of key information

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin corrosion: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)
GLP compliance:
yes (incl. certificate)
Test system:
human skin model
Source species:
other: human derived
Cell type:
non-transformed keratinocytes
Justification for test system used:
Recommended test system in international guidelines (OECD and EC).
Vehicle:
unchanged (no vehicle)
Details on test system:
EpiDerm Skin Model
The model consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multilayered, 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) were cultured on polycarbonate membranes of 10 mm cell culture inserts.
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
The liquid test item was applied undiluted (50 μl) directly on top of the tissue.
Duration of treatment / exposure:
Two tissues were used for a 3-minute exposure to GR-50-3010 and two for a 1-hour
exposure.
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
1
Value:
135
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
2
Value:
127
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Interpretation of results:
GHS criteria not met
Conclusions:
Two tissues were used for a 3-minute exposure to GR-50-3010 and two for a 1-hour exposure.
Executive summary:

GR-50-3010 was checked for color interference in aqueous conditions and possible direct MTT reduction by adding the test item to MTT medium. Because the solutions did not turn blue / purple nor a blue / purple precipitate was observed it was concluded that GR-50-3010

did not interfere with the MTT endpoint.

The mean absorption at 570 nm measured after treatment with GR-50-3010 and controls are presented in Appendix 2, Table 1.

Appendix 3.

Table 2 shows the mean tissue viability obtained after 3-minute and 1-hour treatments with GR-50-3010 compared to the negative control tissues. Skin corrosion is expressed as the remaining cell viability after exposure to the test item. The relative mean tissue viability obtained after the 3-minute and 1-hour treatments with GR-50-3010 compared to the negative control tissues was 135% and 127% respectively. Because the mean relative tissue viability for GR-50-3010 was not below 50% after 3 minutes treatment and not below 15% after 1 hour treatment GR-50-3010 is considered to be not corrosive.

The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the acceptance limits of OECD 431 (lower acceptance limit ≥0.8 and upper acceptance limit 2.8) and the laboratory historical control data range. The mean relative tissue viability following the 1-hour exposure to the positive control was 9%.

In the range of 20 - 100% viability the Coefficient of Variation between tissue replicates was 2.3%, indicating that the test system functioned properly.

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
GLP compliance:
yes (incl. certificate)
Test system:
human skin model
Source species:
other: human derived
Cell type:
non-transformed keratinocytes
Justification for test system used:
In the interest of sound science and animal welfare, a sequential testing strategy is recommended to minimise the need of in vivo testing. One of the validated in vitro skin
irritation tests is the EPISKIN test, which is recommended in international guidelines (e.g. OECD and EC).
Vehicle:
unchanged (no vehicle)
Details on test system:
EPISKIN Small ModelTM (EPISKIN-SMTM, 0.38 cm2, Batch no.: 16-EKIN-047).
This model is a three-dimensional human epidermis model, which consists of adult humanderived epidermal keratinocytes which have been seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. The keratinocytes were cultured for 13 days, which results in a highly differentiated and stratified epidermis model comprising the main basal, supra basal, spinous and granular layers and a functional stratum corneum.
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
The liquid test item was applied undiluted (25 μl) directly on top of the tissue
Duration of treatment / exposure:
15 ± 0.5 minutes
Number of replicates:
The test was performed on a total of 3 tissues per test item together with negative and
positive controls
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
1
Value:
< 50
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Interpretation of results:
Category 2 (irritant) based on GHS criteria
Conclusions:
It is concluded that this test is valid and that GR-50-3010 is irritant in the in vitro skin irritation test under the experimental conditions described in this report.
Executive summary:

In vitro skin irritation test with GR-50-3010 using a human skin model.

This report describes the ability of GR-50-3010 to induce skin irritation on a human three

dimensional epidermal model (EPISKIN Small Model (EPISKIN-SMTM)). The possible skin

irritation potential of GR-50-3010 was tested through topical application for 15 minutes.

The study procedures described in this report were based on the most recent OECD and EC

guidelines.

Batch VE00466805 of GR-50-3010 was a colourless to very slightly yellow liquid.

GR-50-3010 was applied undiluted (25 μl), directly on top of the skin tissue for

15 ± 0.5 minutes. After a 42 hour post-incubation period, determination of the cytotoxic

(irritancy) effect was performed. Cytotoxicity is expressed as the reduction of mitochondrial

dehydrogenase activity measured by formazan production from MTT at the end of the

treatment.

Skin irritation is expressed as the remaining cell viability after exposure to the test item. The

relative mean tissue viability obtained after 15 ± 0.5 minutes treatment with GR-50-3010

compared to the negative control tissues was 4.5%. Since the mean relative tissue viability for

GR-50-3010 was below or equal to 50% after 15 ± 0.5 minutes treatment it is considered to

be irritant.

The positive control had a mean cell viability of 11% after 15 ± 0.5 minutes exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the laboratory historical control data range. The standard deviation value of the percentage viability of three tissues treated identically was less than 3%, indicating that the test systemfunctioned properly.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Species:
other: cow
Details on test animals or tissues and environmental conditions:
Bovine eyes from young cattle were obtained from the
slaughterhouse (Vitelco, 's Hertogenbosch, The Netherlands),
where the eyes were excised by a slaughterhouse employee as
soon as possible after slaughter.
Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent positive control
yes, concurrent negative control
Amount / concentration applied:
The test item was applied as it is (750 μl) directly on top of the corneas
Duration of treatment / exposure:
Corneas were incubated in a horizontal position for 10+/- 1 minutes at 32 +/- 1°C
Duration of post- treatment incubation (in vitro):
Subsequently the corneas were incubated for 120 +/- 10 minutes at 32 +/- 1°C.
Irritation parameter:
in vitro irritation score
Value:
11
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Interpretation of results:
Category 2 (irritating to eyes) based on GHS criteria
Conclusions:
The negative control responses for opacity and permeability were less than the upper limits of the laboratory historical range indicating that the negative control did not induce irritancy on the corneas. The mean in vitro irritancy score of the positive control (Ethanol) was 45 and was within two standard deviations of the current historical positive control mean. It was therefore concluded that the test conditions were adequate and that the test system functioned properly.
GR-50-3010 induced ocular irritation through both endpoints, resulting in a mean in vitro irritancy score of 11 after 10 minutes of treatment.
Executive summary:

GR-50-3010 induced ocular irritation through both endpoints, resulting in a mean in vitro

irritancy score of 11 after 10 minutes of treatment. This is insufficient to classify as Cat 1 based on GHS criteria and is well below the classification threshold. It is also above the threshoold for non-classification. Therefore the substance has been classified as Cat 2 irritant to the eye which is considered sufficiently protective based on current evidence and avoids the need for additional animal data.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for classification or non-classification

GR-50-3010 induced ocular irritation through both endpoints, resulting in a mean in vitro

irritancy score of 11 after 10 minutes of treatment. This is insufficient to classify as Cat 1 based on GHS criteria and is well below the classification threshold. It is also above the threshoold for non-classification. Therefore the substance has been classified as Cat 2 irritant to the eye which is considered sufficiently protective based on current evidence and avoids the need for additional animal data.

GR 50 -3010 was irritating in vitro skin assay but not corrosive leading to recommendation to classify irritant to the skin.