dilithium trimanganese nickel octoxide

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: 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

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2022

Report date:

2022

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Test material

Test material information

impurity 1

Reference substance name:

Dipotassium oxide

EC Number:

235-227-6

EC Name:

Dipotassium oxide

Cas Number:

12136-45-7

Molecular formula:

K2O

IUPAC Name:

Potassium oxide

impurity 2

Reference substance name:

Disodium oxide

EC Number:

215-208-9

EC Name:

Disodium oxide

Cas Number:

1313-59-3

Molecular formula:

Na2O

IUPAC Name:

sodium oxide

impurity 3

Reference substance name:

Lithium sulphate

EC Number:

233-820-4

EC Name:

Lithium sulphate

Cas Number:

10377-48-7

Molecular formula:

H2O4S.2Li

IUPAC Name:

dilithium sulfate

impurity 4

Reference substance name:

Nickel monoxide

EC Number:

215-215-7

EC Name:

Nickel monoxide

Cas Number:

1313-99-1

Molecular formula:

NiO

IUPAC Name:

Nickel (II) oxide

impurity 5

Reference substance name:

dilithium manganese nickel tetraoxide

Cas Number:

128975-24-6

Molecular formula:

Li2MnNiO4

IUPAC Name:

dilithium manganese nickel tetraoxide

Constituent 1

Reference substance name:

dilithium trimanganese nickel octaoxide

EC Number:

663-306-9

Cas Number:

12031-75-3

Molecular formula:

Li2Mn3NiO8

IUPAC Name:

dilithium trimanganese nickel octaoxide

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

Batch no 2021083735

In vitro test system

Test system:

other: Reconstructed Human Epidermis (RHE), EpiDerm is a ready-to-use, highly differentiated 3D tissue model consisting of normal, human-derived epidermal keratinocytes (NHEK) cultured on specially prepared tissue culture inserts.

Source species:

human

Justification for test system used:

In OECD Test Guideline No. 431 EpiDerm™ Skin Corrosivity Test (SCT) (EPI-200 and EPI-212) is referred to as a Validated Reference Method that – based on prevalidation studies, followed by a formal validation study for assessing skin corrosion – could be used for regulatory purposes.

Details on test system:

The reconstructed human epidermal model EpiDerm™ (EPI-200 and EPI-212, MatTek, Ashland, MA, USA and Bratislava, Slovakia) consists of normal human-derived epidermal keratinocytes, which have been cultured to form a multi-layered, 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. A generic description of general and functional conditions that reconstructed human skin models need to comply with can be found in the new OECD Test Guidelines No. 431 In Vitro Skin Corrosion: Reconstructed Human Epidermis (RhE) Test Method and No. 439 In vitro Skin Irritation: Reconstructed Human Epidermis Test Methods.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

First, 25 µL of sterile deionized water was applied to the epidermal surface in order to improve further contact between powder and epidermis. Subsequently, approximately 25 mg of the test item was applied evenly to the epidermal surface. The test item was spread gently on the skin surface with a curved flat spatula without damaging the epidermis. The insert was gently shaken from side to side to ensure that tissue was completely covered by the test item.

Duration of treatment / exposure:

1 hour + 3 min

Number of replicates:

2 replicates of test item, 2 replicates of negative control, 2 replicates of positive control, 2 replicates of colour controls (NSCliving), 2 test item treated killed tissues for non-specific colour control (NSCkilled), 2 killed test item treated tissues and 2 killed negative control treated tissues were used for the MTT evaluation.

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Mean

Value:

96

Negative controls validity:

valid

Positive controls validity:

valid

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In conclusion, in this in vitro skin corrosion test in the EpiDermTM model (OECD 431) with LNMO cathode material the results indicate that the test item is not corrosive. According to the UN GHS classification system, LNMO cathode material has been categorized as “Non-corrosive”.

Executive summary:

Disks of EpiDerm^TM (two units / chemical / incubation time) were treated with the test item and incubated for 60 minutes (+ 3 min) at standard culture condition (37±1 °C in an incubator with 5±1 % CO₂ in a 95±1 % % humidified atmosphere) and for 3 minutes at room temperature. Exposure of test material was terminated by rinsing with 1x DPBS solution. The viability of each disk was assessed by incubating the tissues for 3 hours (±5 min) with MTT solution at 37±1 °C in an incubator with 5±1 % CO₂ in a 95±1 % humidified atmosphere and protected from light. An exception were the additional colour controls (NSC_living and NSC_killed), which wells were filled up with 300 µL assay medium (instead of MTT ready to use solution), the other steps remained the same. The formazan precipitated was then extracted using MTT-100-EXT (extractant solution) and quantified spectrophotometrically.

Potassium hydroxide (KOH) 8N solution and sterile deionized water treated (two units / positive and negative control) epidermis were used as positive and negative controls, respectively in the experiment.

The test item is a possible MTT-reducer, therefore additional controls (two test item treated killed tissues and two negative control treated killed tissues) were used to detect and correct for test item interference with the viability measurement in each exposure time. The mean non-specific MTT reduction (NSMTT) was determined to be 0* % at 60 minutes exposure and 0*% at 3 minutes exposure.

*: The calculated NSMTT was -1% and -4% at 60 and 3 minutes exposure. However, for the calculation of non-specific MTT reduction, small negative numbers are counted as zero, because the reason of the small negative number is a slight difference between the used killed epidermis (biological variability).

The test item has an intrinsic colour (grey/black), therefore two additional test item treated tissues were used for the non-specific OD evaluation (NSC_living).

The test item is a MTT-reducer and has an intrinsic colour (grey/black). To avoid a possible double correction [TOD_TT (MTT and NSC_living)] for colour interference, a third control for non-specific colour in killed tissues (NSC_killed) was performed. However, as the NSC_living % was 2 % and 1 % (below 5 %) at 60 and 3 minutes exposure and the non-specific MTT reduction (NSMTT) was determined to be 0 % in both exposure times, so the NSC_killed was not determined and used during the calculation of true MTT metabolic conversion in each exposure time.

For each treated tissue viability was expressed as a % relative to negative control. The test item is considered to be non-corrosive to skin, if the mean relative viability after 60 minutes of exposure is above or equal to 15 % and the mean relative viability after 3 minutes of exposure is above or equal to 50 % of the negative control.

The test item did not show significantly reduced cell viability in comparison to the negative control after 60 and 3 minutes exposure. The average test item treated tissue mean viability was 96 % at 60 minutes of exposure and the mean viability was 93 % at 3 minutes of exposure.

 

Positive and negative controls showed the expected optical density (OD) in the experiment and cell viability values within acceptable limits. All assay acceptance criteria were met, the experiment was considered to be valid.

In conclusion, in this in vitro skin corrosion test in the EpiDerm^TM model (OECD 431) with LNMO cathode material the results indicate that the test item is not corrosive. According to the UN GHS classification system, LNMO cathode material has been categorized as “Non-corrosive”.