Gallium

Endpoint summary

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

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

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

foreskin from a single donor

Vehicle:

unchanged (no vehicle)

Amount/concentration applied:

Gallium (CAS No. 7440-55-3; EC No. 231-163-8) was warmed to 37°C and was applied as liquid test item topically undiluted to the model skin surface. 30 μL of test item were applied to the skin model and uniformly covered the skin surface with an area of 0.63 cm2.

Duration of treatment / exposure:

The whole exposure period for the used EpiDermTM skin model was 60 minutes. The incubation conditions were 37°C, 5% CO2 and 95% relative humidity for the first 35 minutes followed by 25 minutes at room temperature under a sterile hood.

Duration of post-treatment incubation (if applicable):

At the end of the exposure period, the test item was carefully washed from the skin surface with Dulbecco's phosphate buffered saline (D-PBS). Viability measurements were not performed immediately after the exposure to the test item, but after a post- treatment incubation period of the rinsed tissues in fresh assay medium of 42 hours. This period allows both for recovery from weakly irritant effects and for appearance of clear cytotoxic effects.

Number of replicates:

3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

mean value for 3 replicates

Value:

111.3

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Remarks:

The mean optical density (OD) of 3 negative control tissues was 1.314 and was well within the acceptable range of ≥ 1.0 to ≤ 2.5.

Positive controls validity:

valid

Remarks:

The viability of cells treated with the positive reference item, 5% SDS, was 6.9% of the negative control and fulfilled the acceptance criterion of ≤ 20%.

Remarks on result:

no indication of irritation

Remarks:

The standard deviation determined for all triplicates was below the limit of acceptance of 18%.

Interpretation of results:

GHS criteria not met

Conclusions:

Under the present test conditions, Gallium (CAS No. 7440-55-3; EC No. 231-163- 8) tested at an exposure time of 60 minutes and a 42-hour post-treatment incubation period, was non-cytotoxic and, hence, predicted to be non-irritant to skin in an experiment employing an artificial three-dimensional model of human skin. Hence, the test item did not show irritant properties and is therefore not classified as irritant (UN GHS no category).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not 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:

01/04/2019 - 04/04/2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Batch no 1047000052

Species:

human

Details on test animals or tissues and environmental conditions:

The EpiOcularTM tissue consists of normal, human-derived keratinocytes which have been cultured to form a stratified squamous epithelium similar to that found in the human cor-nea. It consists of highly organized basal cells. These cells are not transformed or trans-fected with genes to induce an extended life span. The EpiOcularTM tissues are cultured in specially prepared cell culture inserts with a porous membrane through which nutrients can pass to the cells. The tissue surface is 0.6 cm2.

EpiOcularTM tissues were procured from MatTek In Vitro Life Science Laboratories, Mlynské Nivy 73, 82105 Bratislava, Slovakia.
Designation of the kit: OCL-200-EIT
Day of delivery: 02. Apr. 2019
Batch no.: 27098

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

50 µL

Duration of treatment / exposure:

28 minutes

Duration of post- treatment incubation (in vitro):

120 minutes

Number of animals or in vitro replicates:

2 replicates

Details on study design:

Preparations

On the day of the start of the experiment, the MTT concentrate was thawed. The MTT concentrate was diluted with assay medium directly before use.
The assay medium was warmed in the water bath to 37 ± 1°C.
6-well-plates were labelled with test item, negative control and positive control and filled with 1 mL assay medium in the appropriate wells. All inserts were inspected for viability and the presence of air bubbles between agarose gel and insert. Viable tissues were transferred in the prepared 6-well-plate and incubated at 37 ± 1 °C, 5 ± 1 % CO2 and ≥ 95% relative humidity for 1 hour.
After the pre-incubation, the medium was replaced and the wells were filled with 1 mL fresh assay medium. All 6-well-plates were incubated at 37 ± 1 °C, 5 ± 1 % CO2 and ≥ 95% relative humidity for 16 hours and 59 minutes.

Exposure and Post-Treatment

After overnight incubation, the tissues were pre-wetted with 20 μL DPBS and then incu-bated at 37 ± 1 °C, 5 ± 1 % CO2 and ≥ 95% relative humidity for 30 minutes. After that, 50 μL of the controls and 50 μL of the test item were applied in duplicate in one- minute-intervals. This was done in such a way that the epithelial surface of the tissue was uniform-ly covered. At the beginning of the experiment (application of negative controls), a stop watch was started. After dosing the last tissue of each plate, the plate was transferred into the incubator for 28 minutes at 37 ± 1 °C, 5 ± 1 % CO2 and ≥ 95% relative humidity.
At the end of the exposure time, the inserts were removed from the plates in one- minute-intervals using sterile forceps and rinsed immediately. The inserts were thoroughly rinsed with DPBS. Then, the tissues were immediately transferred to and immersed in 5 mL of pre-warmed assay medium in a pre-labelled 12-well plate for 12 minutes post soak at room temperature.
After that, each insert was removed from the medium, the medium was decanted off the tissue and the insert was blotted on absorbent material and transferred into the respective well of a pre-labelled 6-well plate containing 1 mL assay medium. For post-treatment incu-bation, the tissues were incubated for 120 minutes at 37 ± 1 °C, 5 ± 1 % CO2 and ≥ 95% relative humidity.
After the post-treatment incubation, the MTT assay was performed.

MTT Assay and Extraction

A 24-well-plate was prepared with 300 μL freshly prepared MTT solution in each well. The tissue inserts were blotted on absorbent material and then transferred into the MTT solu-tion. The plate was incubated for 180 minutes at 37 ± 1 °C, 5 ± 1 % CO2 and ≥ 95% rela-tive humidity.
At last, each insert was thoroughly dried and set into the empty 24-well-plate. Into each well, 2 mL isopropanol were pipetted, taking care to reach the upper rim of the insert. The plate was sealed to avoid evaporation of the solvent and stored in the refrigerator over-night. On the next day, the plate was shaken (120 rpm) for 2 hours at room temperature, protected from light.

Measurement

The inserts were pierced with an injection needle, taking care that all colour was extracted. The inserts were then discarded and the content of each well was thoroughly mixed to achieve homogenisation.
From each well, two replicates with 200 μL solution (each) were pipetted into a 96-well-plate which was read in a plate spectrophotometer at 570 nm.

Irritation parameter:

other: % tissue viability

Run / experiment:

1

Value:

99.3

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation parameter:

other: % tissue viability

Run / experiment:

2

Value:

99.1

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the test, gallium is considered non-eye irritant in the EpiOcularTM Eye Irritation Test.

Executive summary:

Under the conditions of the test, gallium is considered non-eye irritant in the EpiOcularTM Eye Irritation Test.

After treatment with the test item, the mean value of relative tissue viability was reduced to 99.2 %. This value is above the threshold for eye irritation potential (≤ 60%).

All validity criteria were met. The criterion for optical density of the negative control was fulfilled: The OD value was 1.7 (> 0.8 and < 2.5).

The positive control induced a decrease in tissue viability as compared to the negative control to 43.5%. Variation within the replicates of the controls and the test item was acceptable (< 20%).

For these reasons, the result of the test is considered valid.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)