Registration Dossier

Diss Factsheets

Administrative data

Description of key information

Two in vitro tests were available for the testing of skin corrosion and skin irritation. Both tests showed no skin irritant or corrosive effects and were therefore considered for no classification for skin corrosion/irritation.

One in vitro test for eye irritation was performed and showed irritant effects. In addition, one test for eye corrosion was performed, which revealed no corrosive effects. Based on these information, the test item TMA-Tempo needs to be classified for Eye Irrit. 2 according to CLP Regulation (EC) No 1272/2008.

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:
09.03.2018 to 26.04.2018
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)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Justification for test system used:
The EpiDerm Skin Model is a well-established organotypic, three-dimensional model of the human epidermis and is used for in vitro experiments since many years. It is known for its similarity to human skin.
Vehicle:
water
Details on test system:
The test was carried out with the reconstituted three-dimensional human skin model EpiDerm (TM) (MatTek). This skin model consists of normal (non-cancerous), human-derived epidermal keratinocytes (NHEK) which have been cultured to form a multilayered, highly differentiated model of the human epidermis. The NHEK are cultured on chemically modified, collagen-coated cell culture inserts (Millicell (R). The EpiDerm (TM) skin model exhibits in vivo-like morphological and growth characteristics which are uniform and highly reproducible. It consists of organised basal, spinous, granular and cornified layers analogous to those found in vivo.
Control samples:
yes, concurrent vehicle
yes, concurrent positive control
Amount/concentration applied:
25 mg test item and 25 µL H2O
Duration of treatment / exposure:
3 and 60 min
Number of replicates:
2
Irritation / corrosion parameter:
% tissue viability
Remarks:
3 min
Run / experiment:
mean
Value:
92
Vehicle controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Remarks:
60 min
Run / experiment:
mean
Value:
88.6
Vehicle controls validity:
valid
Positive controls validity:
valid
Interpretation of results:
GHS criteria not met
Conclusions:
In this study under the given conditions the test item showed no corrosive effects. The test item is classified as “non-corrosive“.
Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23.04.2018 to 10.07.2018
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
Qualifier:
according to guideline
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Vehicle:
other: Dulbecco’s phosphate buffered saline (DPBS)
Details on test system:
The test was carried out with the reconstituted three-dimensional human skin model EpiDerm (MatTek). This skin model consists of normal human epidermal keratinocytes (NHEK) which have been cultured to form a multilayered, highly differentiated model of the human epidermis. The NHEK are cultured on chemically modified, collagen-coated cell culture inserts (Millicell). The EpiDerm epidermis model exhibits in vivo-like morphological and growth characteristics which are uniform and highly reproducible. It consists of organised basal, spinous and granular layers and a multi-layered stratum corneum analogous to patterns found in vivo.
Control samples:
yes, concurrent vehicle
yes, concurrent positive control
Amount/concentration applied:
25 mg in 25 µL DPBS
Duration of treatment / exposure:
60 min
Duration of post-treatment incubation (if applicable):
42 h
Number of replicates:
3
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
after 60 min treatment and 42 h post-incubation
Value:
75.8
Vehicle controls validity:
valid
Positive controls validity:
valid
Interpretation of results:
GHS criteria not met
Conclusions:
In this study under the given conditions the test item showed no irritant effects. The test item is therefore classified as “non-irritant” in accordance with UN GHS “No Category”.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Referenceopen allclose all

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
06.03.2018 to 16.07.2018
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)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Species:
human
Details on test animals or tissues and environmental conditions:
The test was carried out with the EpiOcular™ reconstructed human cornea-line epithelium (RhCE) model (MatTek). The model consists of normal, human-derived epidermal keratinocytes which have been cultured to form a stratified, highly differentiated squamous epithelium morphologically similar to that found in a human cornea. The EpiOcular™ RhCE tissue construct consists of at least 3 viable layers of cells and a non-keratinized surface, showing a cornea-like structure analogous to that found in vivo.
Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent positive control
yes, concurrent negative control
Amount / concentration applied:
50 mg of test item
Duration of treatment / exposure:
6 h
Duration of post- treatment incubation (in vitro):
18 h
Irritation parameter:
other: mean relative tissue viability [%]
Run / experiment:
1
Value:
13.8
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Irritation parameter:
other: mean relative tissue viability [%]
Run / experiment:
2
Value:
8.9
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid

  1. Name

    Negative Control

    Positive Control

    Test item

    Tissue

    1

    2

    1

    2

    1

    2


    OD570values

    1.665

    1.474

    0.423

    0.393

    0.256

    0.180

    1.645

    1.481

    0.421

    0.392

    0.255

    0.179

    OD570values (blank-corrected)

    1.620

    1.429

    0.378

    0.348

    0.211

    0.135

    1.600

    1.437

    0.377

    0.347

    0.210

    0.134

    mean of the duplicates

    1.610

    1.433

    0.377

    0.347

    0.210

    0.135

    mean OD

    1.521*

    0.362

    0.173

    mean sd OD

    0.125

    0.021

    0.053

    tissue viability [%]

    105.8

    94.2

    24.8

    22.8

    13.8

    8.9

    relative tissue viability difference[%]***


    11.6


    2.0


    5.0

    mean tissue viability [%]

    100.0

    23.8**

    11.3

* Corrected mean OD570 of the negative control corresponds to 100% absolute tissue viability

** Mean relative tissue viability of the positive control is < 50%

*** Relative tissue viability difference of replicate tissues is < 20%

Interpretation of results:
Category 2 (irritating to eyes) based on GHS criteria
Conclusions:
In this study under the given conditions the test item showed irritant effects. The mean relative tissue viability (% negative control) was ≤ 60% (11.3%). The test item is classified as “irritant“ in accordance with UN GHS “Category 1” or “Category 2”. Based on this test alone, a decision on classification into one of the 2 categories is not possible.
Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
05.04.2019 to 17.07.2019
Reliability:
1 (reliable without restriction)
Qualifier:
according to guideline
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)
Version / remarks:
09 October 2017
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Species:
cattle
Strain:
not specified
Details on test animals or tissues and environmental conditions:
eyes were collected from the slaughterhouse, A. Moksel AG, Buchloe, Germany
Vehicle:
physiological saline
Amount / concentration applied:
20% concentration in 0.9% NaCl
Duration of treatment / exposure:
4 hours ± 5 minutes
Number of animals or in vitro replicates:
3 corneas
Irritation parameter:
in vitro irritation score
Run / experiment:
mean value
Value:
1.9
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Irritation parameter:
cornea opacity score
Run / experiment:
mean value
Value:
2.01
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: very slight opacity of the tissue
Interpretation of results:
GHS criteria not met
Conclusions:
According to the evaluation criteria the test item TMA-TEMPO is classified into UN GHS No Category (for irreversible effects on the eye/serious damage to eyes).
Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Skin Corrosion - in vitro Reconstructed Human Epidermis:

This study was performed to analyse the skin corrosivity potential of TMA-Tempo on EpiDerm in vitro human epidermis model according to OECD TG 431.

In the present study TMA-TEMPO was applied topically to the EpiDerm tissue for 3 min and 60 min followed by immediate determination of cytotoxic effects via MTT reduction assay. Corrosivity potential of the test item was predicted from the relative mean tissue viabilities obtained after both treatment periods had been compared to the corresponding negative control tissues. The mean relative tissue viability (% negative control) was ≥ 50% (92.0%) after 3 min treatment and ≥ 15% (88.6%) after 60 min treatment. The test item showed no corrosive effects.

In this study under the given conditions the test item showed no corrosive effects. The test item is classified as “non-corrosive“ in accordance with UN GHS “No Category".

Skin Irritation - in vitro Reconstructed Human Epidermis:

This study was performed to analyse the skin irritatant potential of TMA-Tempo on EpiDerm-Standard Model (EPI-200) human epidermis model according to OECD TG 439.

The test item was applied topically. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT after a 60 min exposure and 42 h post-incubation period and compared to those of the concurrent negative controls. The test item showed no irritant effects. The mean relative tissue viability (% negative control) was > 50% (75.8%) after 60 min treatment and 42 h post-incubation.

In this study under the given conditions the test item showed no irritant effects. The relative mean tissue viability after 60 min of exposure and 42 h post-incubation was > 50%. The test item is therefore classified as “non-irritant” in accordance with UN GHS “No Category”.

Eye Irritation - in vitro EpiOccular

This study was performed to analyse the eye irritating potential of TMA-Tempo on EpiOccular in vitro human corneal epithelium model according to OECD TG 492.

Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT after a 6 h exposure period and 18 h post-treatment period and compared to those of the concurrent negative controls.

The test item showed irritant effects. The mean relative tissue viability (% negative control) was ≤ 60% (11.3%).

In this study under the given conditions the test item showed irritant effects. The test item is classified as “irritant“ in accordance with UN GHS “Category 1” or “Category 2”.

Eye Corrosion - Bovine Corneal Opacity and Permeability Assay

This study was performed to analyse the eye irritancy potential of TMA-Tempo according to OECD TG 437.

The test item was suspended with physiological saline 0.9% NaCl to give a 20% concentration.

An intitial measurement was performed on each of the corneas using the opacitometer. The test item or control were applied to the corneae. After 4 hours incubation

32 ± 1 °C the test item or control were removed, the corneae rinsed, refilled with medium and illuminance measurement was performed. Afterwards permeability of the corneae was determined by measuring spectrophotometrically the transfer of sodium fluorescein. All 3 corneas treated with TMA-Tempo showed slight opacity of the tissue. The mean in vitro irritation score was 1.90 (≤ 3 UN GHS No Category).

According to the evaluation criteria the test item is classified into UN GHS No Category (for irreversible effects on the eye/serious damage to eyes).

Justification for classification or non-classification

Based on the available studies on skin irritation and skin corrosion, the test substance is not classified for skin corrosion/irritation according to CLP Regulation (EC) No. 1272/2008.

Based on the result of eye corrosion testing, the test substance is not classified for eye corrosion according to CLP Regulation (EC) No. 1272/2008. Based on the eye irritating effects in the respective study, the test item is classified for Eye Irrit. 2 according to CLP Regulation (EC) No. 1272/2008.