Registration Dossier

Administrative data

Description of key information

The test item is not considered to be irritating to skin based on the results of the OECD Guideline study 439.

The test item is not considered to be irritating to eye based on the results of the OECD Guideline study 437.

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
Study period:
March 6, 2017 - May 22, 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Version / remarks:
Commission Regulation (EU) No. 640/2012 amending, for the purpose of its adaption to technical progress, Regulations (EC) No. 440/2008 laying down test methods pursuant to Regulation (EC) No. 1907/2006 of the European Parliament and the council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH).
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Version / remarks:
July 28, 2015
Deviations:
no
Qualifier:
according to
Guideline:
other: Skinethic skin irritation test -42bis Standard operating procedure (SOP) 2009
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
No pre-treatment ;The test item was applied neat to the tissues.
Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Cell source:
foreskin from a single donor
Justification for test system used:
standard model
Vehicle:
unchanged (no vehicle)
Remarks:
No vehicle used in this study; The test item was applied neat to the tissues.
Details on test system:
CELL CULTURE
- Supplier: EpiSkin/SkinEthic Laboratories, Lyon, France)
- Source: human keratinocytes cultured on a polycarbonate filter in conditions which permit their terminal differentiation
- Format: 24 well plate
- Batch: 17-RHE-038
- Expires: April 3, 2017

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment: room temperature
- Temperature of post-treatment incubation: 37°C

REMOVAL OF THE TEST MATERIAL AND CONTROL
After the end of the treatment interval, the residual test item was removed immediately by gently rinsing with a minimum volume of 25 mL DPBS using a pipette. Excess DPBS was removed by gently shaking the inserts and blotting the bottom with blotting paper.

DYE BINDING METHOD
- Dye used in the dye-binding assay: MTT
- Spectrophotometer:ELx800, BioTek Instruments GmbH, Bad Friedrichshall, Germany at 570 nm
Control samples:
yes, concurrent negative control
Amount/concentration applied:
TEST MATERIAL: 16 mg of solid test material
NEGATIVE CONTROL: 16 µL (Dulbecco`s Phosphate-Buffered Saline)
POSITIVE CONTROL: 16 µL (5% aqueous solution of sodium dodecyl sulfate in deionised water)
Duration of treatment / exposure:
42 min (± 1 minute)
Duration of post-treatment incubation (if applicable):
42 hours (± 1 hour)
Number of replicates:
3
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
Experiment 1 / Run 1
Value:
96.2
Vehicle controls validity:
not applicable
Remarks:
The test item was applied neat to the tissues
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
OTHER EFFECTS:
- Visible damage on test system: none
- Direct-MTT reduction: none
- Colour interference with MTT: none

ACCEPTANCE OF RESULTS:

Acceptability of the Quality Control Data of the Skin Model with Reference to Historical Batch Data:
The negative control OD values were 1.906, 1.663 and 1.766 and, thus, in the range of ≥0.8 and ≤3.0.

Acceptability of the Positive and Negative Control Data:
After treatment with the positive control (5% aqueous solution of sodium dodecyl sulfate) the mean viability value was 1.4% (standard deviation: 21.4%) and, thus, lower than the historically established threshold of 3.03%.
After treatment with the negative control (DPBS-buffer) the mean OD was 1.778 (standard deviation: 6.9%) and, thus, higher than the historically established threshold of 1.459.
Variability of the Data:

The standard deviation between the three tissues replicates treated with the test item was 9.9% and, thus, ≤18%. The standard deviation between the three tissue replicates of the negative control was 6.9%.
The standard deviation between the three tissues replicates treated with the positive control was 21.4% and, thus, >18%. Due to the very irritant potential of the positive control very low viabilities were obtained. Little differences between very low viabilities resulted in a high standard deviation >18%. However, this does not influence the outcome or validity of the study.


The study met all acceptance criteria.


 Group Time / [min]  Mean OD  Mean Relative viability / [%]
 Negative Control 42  1.778 100 
 Positive Control 42

0.025

1.4

 Test Material

42

1.710

96.2

Interpretation of results:
GHS criteria not met
Remarks:
UN GHS: No Caterory (according to OECD TG 439)
Conclusions:
This study was performed according to GLP and the methods applied are fully compliant with OECD TG 439. Under the conditions of the present study, the test item is not considered to possess an irritant potential to skin (UN GHS: No Category).
Executive summary:

This study was performed according to GLP and the methods applied are fully compliant with OECD TG 439. Under the conditions of the present study, the test item is not considered to possess an irritant potential to skin (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:
Apr 26 - Aug 21, 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
other: OCD 437
Version / remarks:
09 October 2017
Deviations:
no
Qualifier:
according to
Guideline:
EU method B.47 (Bovine corneal opacity and permeability test method for identifying ocular corrosives and severe irritants)
Version / remarks:
Commission Regulation (EU) No. 1152/2010 amending, for the purpose of its adaption to technical progress, Regulation (EC) No. 440/2008 laying down test methods pursuant to Regulation (EC) No. 1907/2006 of the European Parliament and the council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH).
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
PREPARATION OF THE TEST MATERIAL
The test item was prepared as a 20% (w/v) suspension in a 0.9% sodium chloride solution. The stability in the vehicle was not investigated. The test item preparation was administered within <1 hour after preparation.
Vehicle:
physiological saline
Controls:
yes, concurrent vehicle
yes, concurrent positive control
Amount / concentration applied:
Preparation

The test item was prepared as a 20% (w/v) solution in a 0.9% sodium chloride solution. The stability in the vehicle was not investigated. The test item preparation was administered within <1 hour after preparation.

TEST MATERIAL: 750 µL (i.e. 150mg/750µL)
NEGATIVE / VEHICLE CONTROL: 750 µL

Designation: 0.9% sodium chloride solution
Supplier: B. Braun Melsungen AG, Germany
Batch: 16435011
Storage: 2 to 8°C
Released until: Sep 2019

POSITIVE CONTROL: 750 µL

Designation: Art. 814223
Synonym: Imidazole
Supplier: Merck KGaA, Germany
Batch: S6746923
Purity (GC) 99.8% (a/a)
Storage: At room temperature
Released until: Aug 31, 2018



Imidazole was dissolved with 0.9% sodium chloride solution to a concentration of 20% (w/v)
Duration of treatment / exposure:
240 minutes
Number of animals or in vitro replicates:
in vitro: triplicate design
Irritation parameter:
in vitro irritation score
Run / experiment:
Run 1 / Experiment 1
Value:
0
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
ACCEPTANCE OF RESULTS:
After treatment with the negative control (0.9% sodium chloride solution) the calculated IVIS was 0.9 and, thus, within three standard deviations of the current historical mean of the negative control (IVIS: -1.4 – 3.2). After treatment with the positive control (20% Imidazole) the calculated IVIS was 103.8 and, thus, also within two standard deviations of the current historical mean of the positive control (IVIS 80.3 – 133.2). Therefore, the study fulfilled the acceptance criteria.
The resulting classification of the test item in this study is unequivocal and no borderline results were obtained. Therefore, a single testing run composed of three corneas per group was considered sufficient.


Opacity
Permeability
IVIS
per cornea
per group
(mean value)
Standard
deviation
Negative
Control
0.9 % sodium
chloride solution
-0.8
0.001
-0.785
0.9
1.5
1.4
0.001
1.415
2.1
-0.001
2.085
Positive
Control
Imidazole
(20 %)
65.8
1.465
87.775
103.8
13.8
86.6
1.678
111.770
84.5
1.817
111.755
Test Item
-
-1.9
-0.001
-1.915
0.0
2.2
2.4
-0.001
2.385
-0.4
0.003
-0.355


Interpretation of results:
GHS criteria not met
Conclusions:
Under the conditions of the present study, did not show an eye hazard potential. The test item is not requiring classification for eye irritation or serious eye damage (UN GHS: No Category).
Executive summary:

The informtion for this endpoint study record was obtained from an experimental study. The OECD GLP criteria were met and the methods applied are fully compliant with OECD TG 437. Under the conditions of the present study, did not show an eye hazard potential. The test item is not requiring classification for eye irritation or serious eye damage (UN GHS: No Category).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for classification or non-classification

The test item is neither classified for skin irritation/corrosion nor for eye irritation/corrosion according to Regulation (EC) No 1272/2008.