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EC number: 947-684-5 | 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:
- December 2016
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 016
- Report date:
- 2016
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
Test material
- Reference substance name:
- 73138-58-6, 73138-59-7, 73138-60-0
- IUPAC Name:
- 73138-58-6, 73138-59-7, 73138-60-0
- Details on test material:
- - Name of test material (as cited in study report): LICOWAX R 21 S FL
- Physical state: Slightly yellow flakes
- Analytical purity: 99.6 % (w/w)
Fatty acids, tallow,
Guerbet reaction Products 73138-58-6 21% (w/w)
Fatty acids, tallow,
Guerbet reaction Products, Ca salts 73138-59-7 48% (w/w)
Fatty acids, tallow,
Guerbet reaction Products, Na salts 73138-60-0 31% (w/w)
- Lot/batch No.: DEF2084336
- Expiration date of the lot/batch: 2018-11-13
- Storage condition of test material: Room temperature, protected from light in the tightly closed original container
- Solubility : ≤ 0.01 g/L (20 °C)
Constituent 1
In vitro test system
- Test system:
- human skin model
- Source species:
- human
- Details on animal used as source of test system:
- n.a.; in vitro skin model
- Justification for test system used:
- In the interest of sound science and animal welfare, a sequential testing strategy is recommended to minimise 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:
- PRE-TEST
The test item was tested for its ability to direct formazan reduction. This pre-test was done before starting the main study.
- 6 well of 12 well plate was filled with 2 mL of MTT solution (0.3 mg/L).
- To above 6 wells, 10 mg of the test item was added to 3 wells and 10 µL of buffer to other 3 wells and mixed well.
- Incubated the mixture for 3 hours at 37°C protected from light.
Since, no change in the color (blue or purple) of MTT solution was observed, the test item was identified to have no interaction with MTT.
MAIN STUDY
The assay medium supplied with the kits was kept at 2-8°C (refrigerator).
a) Maintenance medium was prewarmed to 37°C before using.
b) EPISKIN Standard Model™ kit was kept at room temperature in the microbiological safety cabinet until next step.
PREPARATION AND PRE INCUBATION (DAY 0)
This step was conducted in a microbiological safety cabinet
a) 3 wells of a 12 well assay plates were filled with pre-warmed maintenance medium (2 mL per well).
b) EPISKIN Standard Model™ kit was opened and epidermis units were transferred into maintenance medium filled wells, using sterile forceps.
Labelled plate lids by replicate treatment order as follows
Negative control Positive control Test item
Rep 1 (NCR1) Rep 1 (PCR1) Rep 1 (5740R1)
Rep 2 (NCR2) Rep 2 (PCR2) Rep 2 (5740R2)
Rep 3 (NCR3) Rep 3 (PCR3) Rep 3 (5740R3)
Key: Rep/R: Replicate; NC: Negative control; PC: Positive control; 5740= study number for test item Licowax R 21 S FL
c) Incubated at 37 °C, 5% CO2, in a > 95% humidified atmosphere for 24 hours.
APPLICATION OF TEST ITEM AND RINSING (DAY 1)
a) Pre-warmed maintenance medium (2 mL per well) was filled taking into account of the 3 wells per replicate. Plate lids were labeled with test-item (3 wells per test item), or negative control (3 wells) or positive control (3 wells).
Topical applications: 15 minutes treatment
b) 10 μL of negative control and positive control, 10 mg of the test item were applied on the top of the epidermis.
c) Followed the defined application order (Rep 1, Rep 2 and Rep 3).
d) After application, plate containing the treated epidermis was closed with lid and placed in the same cabinet for 15 minutes (± 0.5 minute) at room temperature.
For positive control, re-spread after 7 minutes contact time. And each tissue applications were done at 1 minute interval.
End of the treatment and removal of the test chemical; After 15 minutes exposure:
e) Treated epidermal units were removed using forceps, and rinsed thoroughly with 25 mL sterile DPBS by filling and emptying the tissue inserts to remove residual material from the epidermal surface.
f) Epidermal units were placed on an absorbent paper and remaining DPBS were removed from the epidermal surface by gently taping, and swept the epidermal surface with a cotton-bud without damaging the epidermis.
g) Blotted tissue units were transferred to the new maintenance medium pre-filled wells.
Post treatment incubation:42 hours
Incubated the treated and rinsed epidermis at 37°C, 5% CO2, 95% humidified atmosphere for 42 hours.
MTT TEST AFTER THE 42 hours INCUBATION PERIOD
Required number of wells of the assay plate were filled with 2 mL per well of 0.3 mg/mL MTT in assay medium.
Treated units were transferred to the MTT filled wells. Plate was covered with lid and incubated for 3 hours at 37°C, 5% CO2, 95% humidified atmosphere.
FORMAZAN EXTRACTION (DAY 3)
a) 1.5 mL of conic “safelock” type micro tubes were labelled appropriately for test item, vehicle control, positive control and their replicate Number (1 tube/epidermis).
Formazan extraction :
b) Tissue units were placed on absorbent paper to dry the tissues.
c) Using the special biopsy punch, a total biopsy of the epidermis was done.
d) Gently separated the epidermis from the collagen matrix with the aid of forceps, and both parts (epidermis and collagen matrix) were placed into the labelled micro tubes.
e) When all the tissues were punched and ready in the micro tubes, 500 μL of acidic isopropanol per tube was added.
f) Each tube was plugged to avoid evaporation and mixed thoroughly using a vortex mixer.
g) All the tissues were correctly immersed in the solvent and extraction start time for each tissue was noted.
h) The tubes were stored for 4 hours at room temperature (recommended 18 to 23°C) and protected from light.
i) Vortex each tube at the middle of the incubation period.
j) Tissues were removed from each tube extraction end time was noted.
k) Extraction solution was homogenized by pipetting 3 times up and down.
l) From each well, two replicates with 200 μL solution (each) were pipetted into a 96-well-plate which was read in using spectrophotometer at 560 nm. Acidified isopropanol solution was used as blank. - Control samples:
- other: 5% SDS, DPBS
- Amount/concentration applied:
- 10 mg
- Duration of treatment / exposure:
- 15 min
- Duration of post-treatment incubation (if applicable):
- 42 h
- Number of replicates:
- 3
Results and discussion
In vitro
Results
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- mean of three replicates
- Value:
- 93.78
- Vehicle controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Other effects / acceptance of results:
- The negative controls mean OD was found to be 0.726, which is well within the acceptability range. The % viability of tissue treated with the positive control was 24.13% and it was less than 50% of the negative control, which reflects the irritant nature of positive control and the sensitivity of the tissues used in the study.
Applicant's summary and conclusion
- Interpretation of results:
- GHS criteria not met
- Conclusions:
- The % viability of tissue treated with the test item was 93.78% and it was greater than 50% of the negative control. The variation within the replicates was found to be less than 18%. Hence under the conditions of the study the test item was found to be Non Irritant to skin in accordance with UN GHS as specified in the OECD Guideline for the Testing of Chemicals.
- Executive summary:
Test item, Licowax R 21 S FL was applied topically to the EPISKINTMepidermal model (three epidermis units were used per test item, positive and negative controls) for 15 minutes. Exposure to the test item was terminated by rinsing with Dulbecco’s phosphate buffered saline (DPBS). Epidermis was then incubated at 37°C for 42 additional hours. The viability was assessed by incubating the tissues for 3 hours with MTT solution in a 12 well plate (0.3 mg/mL; 2 mL per well). The precipitated formazan was then extracted using acidified isopropanol (0.5 mL) for 4 hours at room temperature, and quantified spectrophotometrically at 560 nm using 96 well plates (200 μL/well).
SDS 5% and DPBS treated epidermis were used as positive (PC) and negative controls (NC) respectively. For each treated tissue the viability was expressed as the % of the negative control tissues (mean).
The negative controls mean OD was found to be 0.726, which is well within the acceptability range. The % viability of tissue treated with the positive control was 24.13% and it was less than 50% of the negative control, which reflects the irritant nature of positive control and the sensitivity of the tissues used in the study.
The % viability of tissue treated with the test item was 93.78% and it was greater than 50% of the negative control. The variation within the replicates was found to be less than 18%. Hence under the conditions of the study the test item was found to beNonIrritantto skin in accordance with UN GHSas specified in the OECD Guideline for the Testing of Chemicals.
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