Single Wall Carbon Nanotubes (SWCNT)

Endpoint summary

Description of key information

A study was performed in accordance with OECD TG 431: the test item was considered to be non-corrosive to the skin.

A study was performed in accordance with OECD TG 439: the test item was classified as non-irritant to skin.

A study was performed in accordance with OECD TG 492: under the experimental conditions reported, the test item possesses an eye irritating potential.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13 July 2016 - 15 July 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Version / remarks:

OECD Guideline for the Testing of Chemicals No. 431 In Vitro Skin Corrosion: Reconstructed Human EpiDermis (RHE) Test Method (28 July 2015).

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

other: epithelial, derived from human skin, and formed into a stratified, cornified epithelium

Vehicle:

unchanged (no vehicle)

Details on test system:

EpiDerm™ Reconstructed Human Epidermis Model Kit
Supplier : MatTek
EpiDermTM Tissues (0.63cm2) lot number : 23345
Assay Medium lot number : 080716ZSB

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

25 mg

Duration of treatment / exposure:

3 Minutes
60 minutes

Duration of post-treatment incubation (if applicable):

3 hours

Number of replicates:

2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minutes exposure

Value:

97.8

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minutes exposure

Value:

97.9

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

The MTT solution containing the test item did not turn blue/purple. However, due to an inconclusive result, corrective procedures were run in parallel on freeze-killed tissues as a precaution.
The results obtained showed that no interference due to direct reduction of MTT occurred and it was therefore considered unnecessary to use the results of the freeze-killed tissues for quantitative correction of results or for reporting purposes.
The mean OD562 for the negative control treated tissues was 1.728 for the 3-Minute exposure period and 1.765 for the 60-Minute exposure period. The negative control acceptance criteria were therefore satisfied.
The relative mean tissue viability for the positive control treated tissues was 5.6% relative to the negative control following the 60-Minute exposure period. The positive control acceptance criterion was therefore satisfied.
In the range 20 to 100% viability the Coefficient of Variation between the two tissue replicates of each treatment group did not exceed 30%. The acceptance criterion was therefore satisfied.

Interpretation of results:

GHS criteria not met

Conclusions:

The test item was considered to be non-corrosive to the skin.

Executive summary:

A study was performed in accordance with OECD TG 431 to evaluate the corrosivity potential of test item using the EpiDerm™ Human Skin Model after treatment periods of 3 and 60 minutes.

Corrosion is directly related to cytotoxicity in the EpiDerm™ tissue. Cytotoxicity is determined by the reduction of MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to formazan by viable cells in the test item treated tissues relative to the corresponding negative control. The results are used to make a prediction of the corrosivity potential of the test item.

Duplicate tissues were treated with the test item for exposure periods of 3 and 60 minutes. Negative and positive control groups were treated for each exposure period. The test item was found to be inconclusive in the MTT direct reduction test therefore additional non-viable tissues were incorporated into the testing for correction purposes as a precaution. At the end of the exposure period the test item was rinsed from each tissue before each tissue was taken for MTT-loading. After MTT loading each tissue was placed in 2 mL Isopropanol for MTT extraction. At the end of the formazan extraction period each well was mixed thoroughly and triplicate 200 mL samples were transferred to the appropriate wells of a pre-labeled 96-well plate. The optical density (OD) was measured at 562 nm (OD562). Data are presented in the form of percentage viability (MTT reduction in the test item treated tissues relative to negative control tissues).

The relative mean viabilities for each treatment group, with negative control set as 100%, were 97.8% after 3 minutes and 97.9% after 60 minutes. The quality criteria required for acceptance of results in the test were satisfied.

The test item was considered to be non-corrosive to the skin.

Reference 2

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

26 July 2016 - 01 August 2016

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

Vehicle:

unchanged (no vehicle)

Details on test system:

EPISKIN™ Reconstructed Human Epidermis Model Kit
Supplier : SkinEthic Laboratories, Lyon, France
EpiSkinTM Tissues (0.38cm2)

Control samples:

yes, concurrent negative control

other: Dulbecco’s Phosphate Buffered Saline (DPBS) with Ca++ and Mg++ - negative control ; Sodium Dodecyl Sulphate (SDS) - positive control

Amount/concentration applied:

Approximately 10 mg (26.3 mg/cm2) of the test item was then applied to the epidermal surface

Duration of treatment / exposure:

15 minutes

Duration of post-treatment incubation (if applicable):

42 hours

Irritation / corrosion parameter:

other:

Remarks:

% formazan production compared to negative control

Run / experiment:

Mean of 3 tissues

Value:

ca. 101.3

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Remarks:

15.1%

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

The relative mean tissue viability for the positive control treated tissues was 15.1% relative to the negative control treated tissues and the standard deviation value of the viability was 1.2%.
The positive control acceptance criteria were therefore satisfied. The mean OD562 for the negative control treated tissues was 0.707 and the standard deviation value of the viability was 10.9%. The negative control acceptance criteria were therefore satisfied.
The standard deviation calculated from individual tissue viabilities of the three identically test item treated tissues was 8.5%. The test item acceptance criterion was therefore satisfied.

Interpretation of results:

other:

Remarks:

The test item was classified as non-irritant.

Conclusions:

The test item was classified as non-irritant. The following classification criteria apply:

EU CLP Not classified for Irritation.
UN GHS Not classified for Irritation (category 3 can not be determined).

Executive summary:

The purpose of this test was to evaluate the skin irritation potential of the test item using the EPISKINTM reconstructed human epidermis model after a treatment period of 15 minutes followed by a post-exposure incubation period of 42 hours. The principle of the assay was based on the measurement of cytotoxicity in reconstructed human epidermal cultures following topical exposure to the test item by means of the colorimetric MTT reduction assay. Cell viability is measured by enzymatic reduction of the yellow MTT tetrazolium salt (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to a blue formazan salt (within the mitochondria of viable cells) in the test item treated tissues relative to the negative controls.

Triplicate tissues were treated with the test item for an exposure period of 15 minutes. The test item produced an inconclusive result in the MTT direct reduction pre-test and therefore additional non-viable tissues were incorporated into the testing for correction purposes as a precaution. At the end of the exposure period each tissue was rinsed before incubating for 42 hours. At the end of the post-exposure incubation period each tissue was taken for MTTloading. The maintenance medium from beneath each tissue was transferred to pre-labeled micro tubes and stored in a freezer for possible inflammatory mediator determination. After MTT-loading a total biopsy of each epidermis was made and placed into micro tubes containing acidified isopropanol for extraction of formazan crystals out of the MTT-loaded tissues. At the end of the formazan extraction period each tube was mixed thoroughly and duplicate 200 μL samples were transferred to the appropriate wells of a pre-labeled 96-well plate. The optical density was measured at 562 nm. Data are presented in the form of percentage viability (MTT reduction in the test item treated tissues relative to negative control tissues).

The relative mean viability of the test item treated tissues was 101.3% after the 15-Minute exposure period and 42-Hours post-exposure incubation period.

The quality criteria required for acceptance of results in the test were satisfied.

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:

27 July 2016 - 29 July 2016

Reliability:

1 (reliable without restriction)

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)

Species:

human

Details on test animals or tissues and environmental conditions:

The EpiOcular™ tissues

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

Approximately 50 mg of the test item (83.3 mg/cm2 according to guideline)

Duration of treatment / exposure:

6 hours

Duration of post- treatment incubation (in vitro):

18 hours

Number of animals or in vitro replicates:

duplicate

Details on study design:

.

Irritation parameter:

other: % formazan production compared to negative control

Run / experiment:

2 tissues

Value:

ca. 26.5

Negative controls validity:

valid

Remarks:

0.8 - 2.5 %

Positive controls validity:

valid

Remarks:

32.6 %

Other effects / acceptance of results:

The negative control OD is > 0.8 and < 2.5 (1.387 and 1.452).
The mean relative viability of the positive control is below 50% of the negative control viability (32.6%).
The difference of viability between the two relating tissues of a single item is < 20% (values between 0.9% to 2.8%) in the same run (for positive and negative control tissues and tissues of single test items). This did not apply to the killed controls (items and negative killed control) and the colorant controls which were calculated as percent values related to the viability of the relating negative control.

Interpretation of results:

Category 2 (irritating to eyes) based on GHS criteria

Conclusions:

Irritating effects were observed following incubation with TUBALL®. Compared with the value of the negative control the relative mean absorption value corresponding to the viability of the tissues decreased below 60% (26.5%).
In conclusion, it can be stated that in this study and under the experimental conditions reported, TUBALL® possesses an eye irritating potential.

Executive summary:

This in vitro study was performed to assess the eye irritation potential of TUBALL^® by means of the Human Cornea Model Test.

Because of the intense black colour of the test item additional tests with freeze killed tissues and with viable tissues were performed.

About 50 mg of the test item and each 50 µL of the controls, respectively, were applied to each of duplicate EpiOcular^™ tissue for 6 hours.

Treatment with the positive control induced a decrease in the mean relative absorbance compared with the negative control to 32.6%, thus the validity of the test system is ensured.

The acceptance criteria were met.

Relevant irritating effects were observed following 6 hours incubation with TUBALL^®. The mean relative absorption value of the tissues corresponding to the cornea viability decreased to 26.5% compared with the value of the negative control (threshold for irritancy: ≤ 60%).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Test item was considered to be irritant to eyes under the conditions of the test.