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Diss Factsheets
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EC number: - | CAS number: -
- 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
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- 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:
- 30 May 2017 - 26 June 2017
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 017
- Report date:
- 2017
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Version / remarks:
- 28 July 2015
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
- Version / remarks:
- 20 July 2012
- Deviations:
- no
- GLP compliance:
- yes
Test material
- Test material form:
- liquid
- Details on test material:
- - Physical appearance: pale yellow liquid
- Storage conditions: at room temperature
Constituent 1
In vitro test system
- Test system:
- human skin model
- Source species:
- human
- Cell type:
- non-transformed keratinocytes
- Cell source:
- skin obtained from plastic surgery from multiple donors
- 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: EPISKIN Small Model(TM)
- Tissue batch number(s): 17-EKIN-025
- Surface: 0.38 cm^2
- Expiration date: 26 June 2017
TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature
- Temperature of post-treatment incubation (if applicable): 36.3 - 37.5°C
REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: tissues were washed with PBS (1 washing step)
- Observable damage in the tissue due to washing: no
MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE: The test substance was checked for possible direct MTT reduction and color interference in a previous skin corrosion test using EpiDerm as a skin model.
- Incubation time: 3 hours
- Measurement method: TECAN Infinite M200 Pro Plate Reader (570 nm)
- MTT concentration: 0.3 mg/mL
NUMBER OF REPLICATE TISSUES: 3 for the test substance, the negative and the positive control each.
CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Because the test item interacted with the MTT endpoint, three killed tissues treated with test item and three killed non treated tissues were used for the cytotoxicity evaluation with MTT.
- Killed tissues: EPISKIN-SMTM, 0.38 cm^2, Lot no.: 17-EKIN-022
- Procedure used to prepare the killed tissues: Living epidermis was transferred to 12 well plates and incubated with 2 ml Milli-Q for 48 ± 1 hours. After incubation, killed epidermis was stored at ≤ -15°C. Killed tissues were
thawed by placing them for 1 hour at room temperature in 12 well plates on 2 ml maintenance medium. Further use of killed tissues was similar to living tissues.
- N. of replicates : 3
- Method of calculation used: the nonspecific MTT reduction (NSMTT) was calculated as follows: %NSMTT = [(ODkt_t+MTT – ODkt_u+MTT)/ mean ODlt_u+MTT] * 100
with (ODkt_u+MTT) = mean OD of the untreated killed tissues; (ODkt_t+MTT) = mean OD of the test item treated killed tissues; (ODlt_u+MTT) = mean OD of the negative control tissues
True tissue viability is calculated as the difference between the living test item treated tissues incubated with MTT medium (ODlt_t+MTT) and the difference between ODkt_t+MTT and ODkt_u+MTT:
OD= ODlt_t+MTT – (ODkt_t+MTT-ODkt_u+MTT)
%Viability = [OD/ mean ODlt_u+MTT] * 100
NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1 single run
CELL VIABILITY MEASUREMENT: After incubation, tissues were dried and incubated with 2 mL MTT-solution for 3 hours. After incubation, epidermis was separated from the collagen matrix and both parts were extracted with 500 μL isopropanol. Tubes were stored refrigerated and protected from light for approximately 68 hours. The amount of extracted formazan was determined spectrophotometrically at 570 nm in duplicate with the TECAN Infinite® M200 Pro Plate Reader.
PREDICTION MODEL / DECISION CRITERIA (see Table 1)
A test item is considered irritant in the skin irritation test if:
The relative mean tissue viability of three individual tissues after 15 minutes of exposure to the test item and 42 hours of post incubation is ≤ 50% of the mean viability of the negative controls.
A test item is considered non-irritant in the in vitro skin irritation test if:
The relative mean tissue viability of three individual tissues after 15 minutes of exposure to the test item and 42 hours of post incubation is > 50% of the mean viability of the negative controls.
ACCEPTABILITY CRITERIA
a) The absolute mean OD570 (optical density at 570 nm) of the three tissues of the negative control should reasonably be within the laboratory historical control data range and the Standard Deviation value (SD) of the % viability should be ≤18.
b) The mean relative tissue viability of the positive control should be ≤50% relative to the negative control and the Standard Deviation value (SD) of the % viability should be ≤18.
c) The SD calculated from individual % tissue viabilities of the three identically treated replicates should be ≤18.
d) The non-specific MTT reduction should be ≤ 30% relative to the negative control OD. - Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- Amount/concentration applied:
- TEST MATERIAL
- Amount applied: 25 μL
NEGATIVE CONTROL
- Amount applied: 25 μL
POSITIVE CONTROL
- Amount applied: 25 μL, re-spread afer 7 minutes of contact time - Duration of treatment / exposure:
- 15 +/- 0.5 minutes
- Duration of post-treatment incubation (if applicable):
- 42 hours; + 3 hours with MTT
- Number of replicates:
- 3
Results and discussion
In vitro
Results
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- Mean of 3 replicates
- Value:
- 78
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks:
- Mean tissue viability: 22%
- Other effects / acceptance of results:
- - OTHER EFFECTS:
- Visible damage on test system: no
- Interaction with the MTT endpoint: yes
- Non-specific reduction of MTT by the test item: 4.80%. The net OD of the treated killed tissues was subtracted from the ODs of the test item treated viable tissues.
ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes, the absolute mean OD570 of the negative control tissues was within the laboratory historical control data range and the SD of the % viability was 5.9%.
- Acceptance criteria met for positive control: yes, the mean relative tissue viability was 22% and the SD was 11%
- Acceptance criteria met for variability between replicate measurements: yes, the standard deviation of the three tissue replicates treated with test substance was 6.8%.
- The non-specific MTT reduction should was < 30% relative to the negative control OD.
- OD values were corrected for background absorption, measured using isopropanol.
- Since the mean relative tissue viability for the test item was above 50% the test item is considered to be non-irritant.
Applicant's summary and conclusion
- Interpretation of results:
- GHS criteria not met
- Conclusions:
- In an in vitro skin irritation test, performed according to OECD guideline 439 and under GLP principles, X-19657 showed to be non-irritant (78% tissue viability after 15 minutes exposure, compared to the negative control). Therefore, the substance is not classified according to the Globally Harmonized System of Classification and Labeling of Chemicals (GHS) of the United Nations.
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
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