Registration Dossier
Registration Dossier
Diss Factsheets
Use of this information is subject to copyright laws and may require the permission of the owner of the information, as described in the ECHA Legal Notice.
EC number: 946-615-6 | 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:
- 25 September to 13 October 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:
- 2018
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:
- adopted 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:
- 23 July 2009
- Deviations:
- no
- Principles of method if other than guideline:
- Not applicable
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- German GLP Compliance Program (inspected on July 13-16, 2015 / Signed on September, 2015)
Test material
- Reference substance name:
- Concrete of honeycomb cells of the bee by hexane extraction
- EC Number:
- 946-615-6
- IUPAC Name:
- Concrete of honeycomb cells of the bee by hexane extraction
- Test material form:
- solid
- Details on test material:
- - Appearance: pale brown solid, wax-like
Constituent 1
- Specific details on test material used for the study:
- - Storage conditions: At room temperature, protected from moisture (Dry area, unopened containers, optimum temp. 11-25 °C / 52-77 °F)
In vitro test system
- Test system:
- human skin model
- Source species:
- human
- Cell type:
- non-transformed keratinocytes
- Justification for test system used:
- Following the REACH top-down strategy, the EPISKIN™ Reconstructed Human Epidermis Model method was used to assess skin irritation as recommended in the OECD test guideline No. 439.
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EPISKIN™ Reconstructed Human Epidermis Model Kit, SkinEthic Laboratories, Lyon, France
- Tissue batch number(s): 17-EKIN-041
- Production date: not reported
- Shipping date: not reported
- Delivery date: 10 October 2017
- Expiry date: 16 October 2017
- Date of initiation of testing: 10 October 2017
TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature
- Temperature of post-treatment incubation (if applicable): 37°C
REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps: After the end of the treatment interval the inserts were immediately removed from the 12-well plate. The tissues were gently rinsed with PBS to remove any residual test material. Excess PBS was removed by gently shaking the inserts and blotting the bottom with blotting paper.
- Observable damage in the tissue due to washing: none reported
- Modifications to validated SOP: none reported
MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 0.3 mg/mL
- Incubation time: 3 hours
- Spectrophotometer: Versamax® Molecular Devices, 85737 Ismaning, Germany, version 4.7.1
- Wavelength: 570 nm
- Filter: without reference filter
- Linear OD range of spectrophotometer: not reported
FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Barrier function: IC50 = 1.9 mg/ml ( ≥ 1.5 mg/ml)
- Morphology: Well-differenciated epidermis consisting of a basal layer, several spinous and granular layers and a thinck stratum corneum
- Contamination: absence of bacteria, fungus and mycoplasma
- Reproducibility: All of the values for the negative and positive control groups fell within the historical ranges of the testing laboratory obtained in the previous ten months. This was taken to show the correct functioning of the test system. Envigo CRS GmbH changed the negative control for the test system “In vitro Skin Irritation Assay using RHE supplied by SkinEthik” from deionized water to PBS. The test is performed at Envigo CRS GmbH a long time prior to OECD 439 guideline, where deionized water or PBS are suggested, and at that time, SkinEthik requested deionized water to be used as negative control in the protocol. All of Envigo CRS GmbH’s historical data are based on deionized water. In the current SkinEthik protocol, PBS is suggested as negative control. Therefore, Envigo CRS GmbH decided to change the negative control to be in accordance with the SkinEthik protocol, but the number of performed assays using PBS as negative control is still too low to issue historical data. SkinEthik confirmed, that use of deionized water in the past is acceptable, since the acceptability ranges for the OD values according to the OECD guideline are met.
CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE: Not a MTT reducer
NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1
PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritating to skin if relative mean tissue viability is ≤ 50% after 15 minutes of exposure.
- The test substance is considered to be non-irritating to skin if relative mean tissue viability is > 50% after 15 minutes of exposure. - Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- Amount/concentration applied:
- TEST MATERIAL
- The test item was used as supplied (undiluted).
- Amount(s) applied (volume or weight with unit): In order to spread approximately 10 ± 5 mg (26 mg/cm2) of the test item onto triplicate EpiSkin™ tissues, 14-15 mg of the test item were weighed to compensate for the test item remained on the applying instrument (curved flat micro spatula). The tissues were wetted with 5 μL of deionised water prior to application of the test substance.
NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 10 μL
- Concentration (if solution): PBS
POSITIVE CONTROL
- Amount(s) applied (volume or weight): 10 μL
- Concentration (if solution): 5% SLS solution in deionised water - Duration of treatment / exposure:
- 15 minutes
- Duration of post-treatment incubation (if applicable):
- 42 hours
- Number of replicates:
- Triplicate tissues for test substance, negative and positive controls
Results and discussion
In vitro
Results
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 15 minute exposure period and 42 h post-exposure incubation period
- Value:
- 96.1
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Remarks:
- 100%
- Positive controls validity:
- valid
- Remarks:
- 2.7%
- Remarks on result:
- no indication of irritation
- Other effects / acceptance of results:
- - OTHER EFFECTS:
- Visible damage on test system: none reported
- Direct-MTT reduction: no
- Colour interference with MTT: no
DEMONSTRATION OF TECHNICAL PROFICIENCY: yes
ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The mean OD570 for the negative control treated tissues was 1.149 and the standard deviation value of the viability was 9.6%.
- Acceptance criteria met for positive control: The relative mean tissue viability for the positive control treated tissues was 2.7% relative to the negative control treated tissues and the standard deviation value of the viability was 23.3%. Therefore the relative Standard deviation of the positive control was greater than 18 %. However, this can be explained by the low OD values measured which implies higher relative standard variations. Moreover the values are lower than the historical positive control data of the laboratory and the results are not bordeline, therefore the relative standard deviation calculated for positive control tissues is not considered to affect the overall conclusion of the study.
- Acceptance criteria met for variability between replicate measurements: The standard deviation calculated from individual tissue viabilities of the three identically test item treated tissues was 11.5%.
- Reference to historical values: All of the values for the negative and positive control groups fell within the historical ranges of the testing laboratory obtained in the previous 10 months. This was taken to show the correct functioning of the test system.
Any other information on results incl. tables
Table 7.3.1/1: EpiSkin™ results
Test Group |
Mean Absorbance 570 nm |
Mean Absorbance of 3 Tissues |
Relative Viability [%] Tissue 1, 2, 3* |
Relative Standard Deviation [%] |
Mean Rel. Viability [%]** |
Negative Control |
1.255 |
1.149 |
109.2 |
9.6 |
100.0 |
1.034 |
90.0 |
||||
1.158 |
100.8 |
||||
Positive Control |
0.040 |
0.031 |
3.5 |
23.3 |
2.7 |
0.028 |
2.4 |
||||
0.026 |
2.3 |
||||
Test Item |
1.116 |
1.104 |
97.1 |
3.5 |
96.1 |
1.062 |
92.4 |
||||
1.135 |
98.8 |
* Relative viability [rounded values]
** Mean relative viability [rounded values]
Applicant's summary and conclusion
- Interpretation of results:
- GHS criteria not met
- Conclusions:
- Under the experimental conditions of this study, the test substance is not classified for skin irritation according to Regulation (EC) No. 1272/2008 (CLP) and to the GHS.
- Executive summary:
An in vitro skin irritation study was performed according to the OECD Guideline 439 and in compliance with GLP, using the EPISKINTM reconstructed human epidermis model.
Triplicate tissues were treated with the test item for an exposure period of 15 minutes. At the end of the exposure period each tissue was rinsed before incubating for 42 hours. At the end of the post‑exposure incubation period each tissue was taken for MTT-loading. The maintenance medium from beneath each tissue was transferred to pre‑labeled micro tubes and stored in a freezer for possible inflammatory mediator determination. After MTT-loading a total biopsy of each epidermis was made and placed into plastic vials containing acidified isopropanol for extraction of formazan crystals out of the MTT‑loaded tissues. At the end of the formazan extraction period each tube was mixed thoroughly and duplicate 200 µL samples were transferred to the appropriate wells of a pre‑labeled 96‑well plate. The optical density was measured at 570 nm.
The relative mean viability of the test item treated tissues was 96.1% after the 15‑Minute exposure period and 42‑Hours post‑exposure incubation period.
The relative mean tissue viability for the positive control treated tissues was 2.7% relative to the negative control treated tissues and the standard deviation value of the viability was 23.3%. Therefore the relative Standard deviation of the positive control was greater than 18 %. However, this can be explained by the low OD values measured which implies higher relative standard variations. Moreover the values are lower than the historical positive control data of the laboratory and the results are not bordeline, therefore the relative standard deviation calculated for positive control tissues is not considered to affect the overall conclusion of the study.
The mean OD570 for the negative control treated tissues was 1.149 and the standard deviation value of the viability was 9.6%. The negative control acceptance criteria were therefore satisfied.
The standard deviation calculated from individual tissue viabilities of the three identically test item treated tissues was 3.5%. The test item acceptance criterion was therefore satisfied.
All of the values for the negative and positive control groups fell within the historical ranges of the testing laboratory obtained in the previous ten months. This was taken to show the correct functioning of the test system.
Under the experimental conditions of this study, the test substance is not classified for skin irritation according to Regulation (EC) No. 1272/2008 (CLP) and to the GHS.
This study is considered as acceptable and satisfies the requirement for skin irritation endpoint.
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.
Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.
