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EC number: 232-001-9 | CAS number: 7783-49-5
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
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- Endpoint summary
- Stability
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- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data

Skin irritation / corrosion
Administrative data
- Endpoint:
- skin irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2018-01-29 to 2018-03-26
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 018
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Version / remarks:
- 2015-07-28
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- signed 2015-09-14
Test material
- Reference substance name:
- Zinc fluoride
- EC Number:
- 232-001-9
- EC Name:
- Zinc fluoride
- Cas Number:
- 7783-49-5
- Molecular formula:
- ZnF2
- IUPAC Name:
- zinc fluoride
- Test material form:
- solid
- Details on test material:
- - State of aggregation: white solid
Constituent 1
- Specific details on test material used for the study:
- STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature. Keep container tightly sealed. Protect from humidity and water
In vitro test system
- Test system:
- human skin model
- Source species:
- human
- Cell type:
- other: normal, human-derived epidermal keratinocytes
- Cell source:
- other: humans
- Source strain:
- not specified
- Details on animal used as source of test system:
- not applicable
- Justification for test system used:
- In an international prevalidation study performed by ECVAM, the in vitro skin irritation test using the human skin model EpiDerm™ and measurement of cell viability by dehydrogenase conversion of MTT into a blue formazan salt have turned out as a sufficiently promising predictor for skin irritancy potential.
- Vehicle:
- other: Dulbecco's phosphate buffered saline
- Details on test system:
- RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ skin model (source: MatTek Corporation, 82105 Bratislava, Slovakia)
- Tissue lot number: 25882
- Delivery date: 2018-02-21
TEMPERATURE USED FOR TEST SYSTEM
- Temperature of pre-incubation: 37 ± 1.5 °C (about 60 minues)
- Temperature used during treatment / exposure: 37 ± 1.5 °C for 35 minutes, room temperature for 25 minutes
- Temperature of post-treatment incubation: 37 ± 1.5 °C
REMOVAL OF TEST MATERIAL AND CONTROLS
After the end of the treatment interval the inserts were rinsed with DPBS at least 15 times in order to remove any residual test material.
After the rinsing the inserts were submerged in DPBS at least three times. Afterwards the inserts were again rinsed with DPBS. The tissues were then transferred into plates with assay medium. Tissues were incubated for about 24 hours at 37 ± 1.5 °C, 5 ± 0.5 % CO2. After incubation medium will be changed (pre-warmed fresh medium). Thereafter tissues were incubated for another approximately 18 hours at 37 ± 1.5 °C, 5 ± 0.5 % CO2. The complete incubation time was 42 hours.
MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL (300 µL/ well)
- Incubation time with MTT: 3 hours
- Extraction of Formazan: after the incubation period, the tissues were rinsed three times with DPBS. The tissues were transferred into new plates containing extractant solution (isopropanol) in each well ensuring that the tissues were completely covered and the plate was sealed to inhibit the isopropanol evaporation. The formazan salt was extracted for about 2.5 hours while shaking at room temperature.
After the extraction period was completed, the inserts were pierced with an injection needle to allow the extract to run into the well from which the insert was taken and the insert was discarded. The 24-well plates were placed on a shaker for 15 minutes until the solution was homogeneous in colour.
Per each tissue, 3 × 200 μL aliquots of the blue formazan solution were transferred into a 96-well flat bottom microtiter plate. The optical density was determined with a microplate reader. Mean values were calculated from the 3 wells per tissue.
- Spectrophotometer: Versamax® Molecular Devices
- Wavelength: 570 nm
TEST FOR COLOUR INTERFERENCE
Before the test started, functional check for colour interference was performed. 25 ± 2 mg of the test item were added to deionised water. The mixture was incubated in the incubator (37 ± 1.5 °C, 5 ± 0.5 % CO2) for 60 minutes. At the end of the exposure time, the mixture was shaken and the presence and intensity of the staining (if any) was evaluated.
TEST FOR DIRECT MTT REDUCTION
The test item was evaluated for its potential to interfere with MTT assay. To test if a test item directly reduces MTT, 25 ± 2 mg of the test item were added to 1 mL of the MTT-solution (1mg/mL) and was incubated in the incubator (37 ± 1.5 °C, 5 ± 0.5% CO2) for 60 minutes. Untreated MTT medium was used as control.
FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: tissues pass analysis for tissue viability
- Barrier function: tissues pass analysis for tissue functionality
- Morphology: presence of a functional stratum corneum, a viable basal cell layer, and intermediate spinous and granular layers
- Contamination: absence of bacteria, yeast, and other fungi (long term antibiotic, antimycotic free culture) as well as absence of HIV1- virus, Hepatitis B virus, and Hepatitis C virus
Please also refer to the field "Attached background material" below.
PREDICTION MODEL / DECISION CRITERIA
The mean optical density (OD) of the three negative control tissues was calculated. This value corresponds to 100% tissue viability in the current test. For each individual tissue treated with the test item or the positive control the individual relative tissue viability is calculated according to the following formula: relative viability(%) = (mean OD test item or positive control/ mean OD of negative control) x 100
For the test item and the positive control, the mean relative viability ± relative standard deviation of the three individual tissues was calculated and used for classification according to the following prediction model: if the mean relative tissue viability of three individual tissues is less or equal to 50% of the negative control, the test item needs to be classified and labeled for its skin irritation potential: Category 2 – irritant, H315 according to Regulation (EC) No 1272/2008. - Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- Amount/concentration applied:
- TEST MATERIAL
- Amount(s) applied (volume or weight with unit): approx. 25 mg of the test item, wetted with vehicle
VEHICLE
- Amount(s) applied (volume or weight with unit): 25 µL DPBS
NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 µL DPBS
POSITIVE CONTROL
- Amount(s) applied (volume or weight): 30 µL of a 5% Sodium Lauryl Sulfate (SLS) solution - Duration of treatment / exposure:
- 60 minutes
- Duration of post-treatment incubation (if applicable):
- 42 hours
- Number of replicates:
- triplicates
Results and discussion
In vitro
Results
- Irritation / corrosion parameter:
- % tissue viability
- Remarks:
- (mean)
- Value:
- 0.8
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Other effects / acceptance of results:
- - OTHER EFFECTS:
- Colour interference with MTT: The test item did not reduce MTT (test for direct MTT reduction), and it did not change colour when mixed with deionised water (test for colour interference).
ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: After treatment with the negative control the absorbance values were well within the required range of the acceptability criterion of mean OD 0.8 and ≤ 2.8 for the 60 minutes treatment interval, thus assuring the quality of the tissues
- Acceptance criteria met for positive control: Treatment with the positive control induced a sufficient decrease in the relative absorbance as compared to the negative control for the 60 minutes treatment interval, and thus assuring the validity of the test system.
- Acceptance criteria met for variability between replicate measurements: Criterion 1 (negative control): The absolute OD 570 nm of the negative control tissues in the MTT test is an indicator of tissue viability obtained after the shipping and storing procedure and under specific conditions of the assay. Tissue viability is meeting the acceptance criterion if the mean OD570 of the negative control tissues is 0.8 and ≤ 2.8 in accordance with OECD TG 439.
Criterion 2 (positive control): An assay is meeting the acceptance criterion if mean relative tissue viability of the positive control is 20%.
Criterion 3: Standard deviation: The SD of 3 concurrently tested tissue replicates should be < 18.
Criterion 4: OD values should not be below historically established boundaries.
Concurrent negative controls (NC) and positive controls (PC) will be used in each run to demonstrate that viability (with the NC), barrier function and resulting tissue sensitivity (with the PC) of the tissues are within a defined historical acceptance range.
Historical data (see annex 1) and the quality certificate of the supplier of the test kit (see annex 3) demonstrating its robustness of the test system, including quality control data (determined by MatTek Corporation, 82105 Bratislava, Slovakia) of the respective EpiDermTM lot. According to the OECD TG 439, the acceptance limit of the ET50 should be between 4.8 hours and 8.7 hours after treatment with 1% Triton X-100 (QC batch release criteria)
Any other information on results incl. tables
Results after treatment with Zinc fluoride and the controls:
Treatment Group |
Tissue No. |
OD 570 nm |
OD 570 nm |
OD 570 nm |
Mean OD of 3 Wells |
Mean OD of 3 Wells blank corrected |
Mean OD of 3 tissues blank corrected |
Rel. Viability [%] Tissue |
Standard Deviation |
Mean Rel. Viability [%]** |
Blank |
|
0.038 |
0.038 |
0.037 |
0.037 |
|
|
|
|
|
Negative Control |
1 |
1.707 |
1.603 |
1.613 |
1.641 |
1.603 |
1.706 |
94.0 |
5.4 |
100.0 |
2 |
1.796 |
1.776 |
1.746 |
1.773 |
1.735 |
101.7 |
||||
3 |
1.862 |
1.809 |
1.782 |
1.817 |
1.780 |
104.3 |
||||
Positive Control |
1 |
0.080 |
0.079 |
0.078 |
0.079 |
0.042 |
0.047 |
2.5 |
0.4 |
2.8 |
2 |
0.093 |
0.093 |
0.094 |
0.093 |
0.056 |
3.3 |
||||
3 |
0.082 |
0.082 |
0.079 |
0.081 |
0.043 |
2.5 |
||||
Test Item |
1 |
0.055 |
0.057 |
0.056 |
0.056 |
0.019 |
0.014 |
1.1 |
0.2 |
0.8 |
2 |
0.047 |
0.048 |
0.048 |
0.048 |
0.010 |
0.6 |
||||
3 |
0.050 |
0.051 |
0.052 |
0.051 |
0.014 |
0.8 |
* Relative viability [rounded values]:
** Mean relative viability [rounded values]:
Historical data:
Positive Control; OD at 570 nm after |
Negative Control OD at 570 nm |
|||||||
MeanViability |
4.28% |
Mean Absorption |
1.66 |
|||||
Standard Deviation |
1.00 p.p. |
Standard Deviation |
0.20 |
|||||
Rel. Standard Deviation |
23.44% |
Rel. Standard Deviation |
11.96% |
|||||
Range of Viabilities |
2.24%—6.19% |
Range of Absorbance* |
1.28—2.00 |
|||||
Mean Absorption |
0.07 |
* should be 0.8—2.8 (OECD 439) |
||||||
Standard Deviation |
0.02 |
|||||||
Rel. Standard Deviation |
25.96% |
|||||||
Range of Absorbance |
0.03—0.11 |
|||||||
Data of 36 sets of controls shared between 147 studies performed from January 2017 until January 2018. (p.p.—percentage points) |
Applicant's summary and conclusion
- Interpretation of results:
- GHS criteria not met
- Conclusions:
- A study on skin irritation in accordance with OECD 429 was performed with zinc difluoride resulting in C&L as skin irritant Cat. 2 according to Regulation (EC) No 1272/2008 and its subsequent adaptations.
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