Naphthenic acids, lithium salts

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because the substance is classified as corrosive to the skin

Cross-reference

Reason / purpose for cross-reference:

data waiving: supporting information

Reference

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 - 18 August 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: Method B.40bis

Version / remarks:

Commision Regulation (EC) No 440/2008, of 30 May 2008, laying down test methods pursuant to Regulation (EC) No 1907/2006, of 18 December 2006, of the European Parliament and of the Council on the Regulation, Evaluation, Authorisation and Restriction of Chemicals (REACH)

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

28 July 2015

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

- Batch No.of test material: A065/99
- Expiration date of the lot/batch: 1 July 2019
- Storage: Room temperature in the dark

Test system:

human skin model

Source species:

human

Cell type:

other: EpiDerm™ Reconstructed Human Epidermis Model Kit. The target cells are epithelial, derived from human skin, and formed into a stratified, cornified epithelium.

Cell source:

other: EpiDerm™ Reconstructed Human Epidermis Model Kit

Source strain:

other: EpiDerm™ Reconstructed Human Epidermis Model Kit

Details on animal used as source of test system:

- Supplier: MatTek
- Date received: 15 August 2017
- EpiDermTM Tissues (0.63cm2) lot number: 25837
- Assay Medium lot number: 081017ALA
- Storage: Upon receipt of the EpidermTM tissues, the sealed 24-well plate was stored in a refrigerator until use.

Justification for test system used:

In the interest of sound science and animal welfare, a sequential testing strategy is recommended to minimize the need of in vivo testing. One of the validated in vitro skin irritation tests is the EPISKIN test, which is recommended in international guidelines (e.g. OECD and EC).

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ Reconstructed Human Epidermis Model Kit
- Tissue batch number(s): 25837
- Production date: Not specified
- Shipping date: Not specified
- Delivery date: 15 August 2017
- Date of initiation of testing: 16 August 2017
- Method: The corrosivity potential of the test item was predicted from the relative mean tissue viabilities obtained after the 3 and 60-Minute exposure periods, compared to the mean of the negative control tissues (n=2) treated with sterile distilled water. The relative mean viabilities were calculated in the following way: Relative mean viability (%) = mean OD570 of test item / mean OD570 of negative control x 100

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

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: Rinsing was achieved by filling and emptying each tissue under a constant soft stream of DPBS to gently remove any residual test item. Excess DPBS was removed by blotting the bottom of the tissue insert with tissue paper.

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA: Not specified

NUMBER OF REPLICATE TISSUES: 2

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 0.1 mg/mL
- Incubation time: 3 hours
- Spectrophotometer: Labtech LT- 4500 microplate reader.
- Wavelength: 570 nm
- Results and implications: The test item was found to produce a colored solution which may interfere with the MTT endpoint. Therefore, color correction tissues were incorporated into the test to correct for this possibility. These tissues were treated identically to the tissues of the main test with the exception of being placed into assay medium for three hours post-exposure instead of MTT. Two tissues were dosed with the test item and two tissues remained untreated for each exposure period.

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Tissues used: EpiDerm™ Reconstructed Human Epidermis Model Kit
- No. of replicates: 2
- Method of calculation used: Relative mean viabilities
- Results: Negligible colour interference occurred and it was therefore considered unnecessary to use the results of the colour correction tissues for quantitative correction of results or for reporting purposes. The MTT solution containing the test item did not turn blue/purple and this was taken to indicate the test item did not reduce MTT.

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 2

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be corrosive to skin if the viability after 3 minutes exposure is less than 50%, or if the viability after 3 minutes exposure is greater than or equal to 50 % and the viability after 1 hour exposure is less than 15%.
- The test substance is considered to be non-corrosive to skin if the viability after 3 minutes exposure is greater than or equal to 50% and the viability after 1 hour exposure is greater than or equal to 15%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied: 0.05 mL of the test item

VEHICLE
- Amount(s) applied: None

NEGATIVE CONTROL
- Amount applied: 50 μL of sterile distilled water

POSITIVE CONTROL
- Amount applied: 50 μL of 8.0 N Potassium Hydroxide

Duration of treatment / exposure:

3 and 60 minutes

Duration of post-treatment incubation (if applicable):

3 hours

Number of replicates:

2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minute

Value:

95.5

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minute

Value:

13.6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- Acceptability criteria: The mean OD570 for the negative control treated tissues was 1.528 for the 3-Minute exposure period and 1.441 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 3.1% 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.
- Conclusion: The test item was considered to be corrosive: UN GHS H314 Combination of sub-categories 1B and 1C.

Interpretation of results:

Category 1 (corrosive) based on GHS criteria

Conclusions:

Naphthenic acids, lithium salt was considered to be corrosive: UN GHS H314 Combination of sub-categories 1B and 1C.

Executive summary:

Naphthenic acids, lithium salts is considered to be skin corrosive using the EPIDERM (TM) human epidermis model. The skin corrosion potential of Naphthenic acids, lithium salts was evaluated using EPIDERM (TM) reconstructed human epidermis model after a treatment period of 3 and 60 minutes followed by a post-exposure period of 3 hours using a colourimetric MTT reduction assay following Method B.40bis of Commission Regulation (EC) No 440/2008 in an experimental proprietary study (Envigo 2018). The quality criteria required for the acceptance of results in the test were satisfied and the study is considered relevant and reliable for use.