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Diss Factsheets
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EC number: 939-273-4 | 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
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Skin irritation / corrosion
Administrative data
- Endpoint:
- skin irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- from April 20, 2012 to June 18, 2012
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: GLP compliant with international guidelines
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 012
- Report date:
- 2012
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
Test material
- Reference substance name:
- Paraffin waxes and Hydrocarbon waxes C14-17, chloro, sulfochlorinated, low sulphonated, saponified
- EC Number:
- 939-273-4
- Molecular formula:
- CnHxCly(SO3Na)z • n= 14-17 • z < 0,5 • y = 0 ÷ 2,7
- IUPAC Name:
- Paraffin waxes and Hydrocarbon waxes C14-17, chloro, sulfochlorinated, low sulphonated, saponified
- Test material form:
- liquid: viscous
Constituent 1
In vitro test system
- Test system:
- human skin model
- Source species:
- human
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- - Model used: SkinEthic reconstructed human tissue model EPISKIN
- Source: SkinEthic Laboratories
- Details: The test site consists of an airlifted, living, multi-layered tissue construct, produced in polycarbonate inserts in serum-free and chemically defined medium, featuring normal ultra-structure and functionality equivalent to human tissue in vivo.
Normal human keratinocytes are used to reconstruct the epithelium. Multiple layers of viable epithelial cells (basal layer, stratum spinosum, stratum granulosum) should be present under a functional stratum corneum. Stratum corneum should be multilayered containing the essential lipid profile to produce a functional barrier with robustness to resist rapid penetration of cytotoxic marker chemicals.
- Storage: According to the supplier procedure, the test system was shipped on Monday and received on Tuesday. At arrival, the plate was opened under a sterile airflow and each insert, containing the epidermal tissue, was carefully taken out and placed in a 12-well plate (supplied) in which each well had previously been filled with 2 ml/well SkinEthic Maintenance Medium.
- Pre-treatment incubation period: Culture dishes were placed in the incubator at 37°C, 5% CO2 and saturated humidity for approximately 24 hours.
Preliminary test
Direct MTT reduction test (Step 1): Non-specific reduction of MTT was evaluated as follows:
two mL of MTT Ready-to-use Solution was incubated with 20 mg of test item at 37°C, 5% CO2 and saturated humidity for 3 hours protected from light, simulating test conditions. Observation of blue or purple appearance of the solution at the end of the incubation time was carried out.
Colouring potential test (Step 2): Chemicals' colouring potential was assessed for potential interaction with the test system.
10 mg of the test item was added to 90 µL of distilled water in a transparent tube and the resulting solution/suspension mixed by using a vortex for 15 minutes.
Colouring of the solution at the end of the incubation time was evaluated.
Sample Test System Treatment Amount/well Number of replicates Sample code
Negative
control Live tissue D-PBS 20 µL 3 N1-N3
Positive
control Live tissue 5% SDS in water 20 µL 3 P1-P3
Test item Live tissue PARAFFIN WAXES
AND HYDROCARBON
WAXES, CHLORO,
SULFOCHLORINATED
SAPONIFIED (C14-C17)
SSP-SAMPLE1 20 µL* 3 F1-F3
*:Even if the test item was a dense cream, it was possible to measure the amount in volume (differently from what carried out in the preliminary phase) using a small syringe of adeguate capaci.
calculation
BLANK negative control positive control test item
0.048 0.831 0.085 0.121
0.054 0.82 0.065 0.112
0.053 0.868 0.07 0.06
0.054 0.92 0.064 0.062
0.049 0.829 0.084 0.037
0.047 0.882 0.074 0.066
Mean 0.051 0.858 0.074 0.076
SD 0.003 0.039 0.009 0.033
CV 5.90% 4.60% 12.20% 43.40%
MTT ASSAY:
The tissue insert and controls were incubated with 2 mL well of MTT ready-to-use solution for approximately 3 hours at 37°C, 5% CO2 and saturated humidity.
At the end of the incubation period, tissues were placed on absorbent paper to dry. A total biopsy was carried out by means of a biopsy punch to allow biopsies of the same dimensions.
The epidermis were separated from the collagen matrix and both placed in a microtube prefilled with 500 µL of acidic isopropanol.
Tubes were mixed by vortexing and preserved for approximately 3 days at 4°C to allow formazan extraction.
At the end of the extraction period, debris were eliminated by short centrifugation of the tubes (approximately 14000 rpm for 2 minutes) and aliquots of 200 µL from each sample were read in duplicate for their absorbance at 595 nm. OD values were recorded. Six aliquots (200 µL) of acidic isopropanol were analyzed and used as blank.
After appropriate blank subtractions and/or corrections for the background controls, means, standard deviations, coefficients of variation, mean relative viability values (percentage relative to the negative control) will be calculated.
REMOVAL OF TEST SUBSTANCE
- Washing: At the end of the exposure, the test item was mechanically removed from the surface and each tissue was rinsed three times with approximately 25 mL of sterile D-PBS filling and empting the tissue insert.
The excess liquid was carefully removed and the sample transferred in new wells pre-filled with 2 mL/well of maintenance medium.
- Other: MEDIA
Dulbecco's Phosphate buffered saline (D-PBS)
Sterile water
MTT Reagent
MTT Stock Solution
MTT Ready-to-use Solution
Acidic Isopropanol
(0.04 N HCl in isopropanol)
Positive control item: 5% (w/v) sodium dodecyl sulphate (SDS) solution, obtained by 1:1 dilution in sterile water (Baxter, batch 11K0903) of a sterile commercial 10% (w/v) SDS solution in water (SIGMA, batch 088K8703)
Negative control item: D-PBS (GIBCO, batch 1098734). - Control samples:
- no
- Amount/concentration applied:
- 20 µL/ epidermis unit
- Duration of treatment / exposure:
- 15 minutes
- Duration of post-treatment incubation (if applicable):
- 42 hours recovery period
- Number of replicates:
- 3
Results and discussion
In vitro
Results
- Irritation / corrosion parameter:
- % tissue viability
- Value:
- ca. 3.1
- Vehicle controls validity:
- not examined
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Other effects / acceptance of results:
- validity criteria:
Negative controls: OD values of the negative control samples 0.600, CV% ≤ 18.
Positive controls: mean viability expressed as percentage of the negative control ≤ 40% and CV% ≤ 18.
Test item data acceptance: CV% ≤ 18.
Applicant's summary and conclusion
- Interpretation of results:
- other: Category 2 (irritant) based on CLP criteria
- Conclusions:
- PARAFFIN WAXES AND HYDROCARBON WAXES, CHLORO, SULFOCHLORINATED SAPONIFIED (C14-C17) SSP-SAMPLE 1 has been judged as irritant by in vitro treatment.
- Executive summary:
The potential of the test item PARAFFIN WAXES AND HYDROCARBON WAXES, CHLORO, SULFOCHLORINATED SAPONIFIED (C14-C17) SSP-SAMPLE l to be irritant to the skin was investigated through an in vitro skin irritation study using a commercial reconstructed human epidermis (RhE) model named EPISKINTM
The test item, as well as controls, were tested for their ability to impair cell viability after an exposure period of 15 minutes followed by a 42 hour recovery period. The final endpoint of the assay is the colorimetric measurement of MTT reduction (blue formazan salt) in the test system being this reaction an index of cell viability.
Before the Main Assay, a preliminary test was carried out to assay the compatibility of the test item with the test system. In particular, the test item was assayed for the ability of reducing MTT and of colouring water per se.
No interaction was recorded between the test item and MTT in test conditions similar to those of the Main Assay. Moreover, no colouring potential of the test item in contact with water was recorded. Thus, no additional control was added in the main phase for the evaluation of non specific coloration which may influence evaluation of results.
In the Main Assay, the test item was applied as supplied in three replicates at the treatment level of 20 µL/epidermis unit each measuring 0.38 cm^2 (treatment level: 53 µL/cm^2). Positive and negative controls [a 5% (w/v) sodium dodecyl sulphate solution in water and Dulbecco's phosphate buffered saline (D-PBS)] were concurrently tested, in the same number of replicates and test conditions at the treatment level of 20 µL/ epidermis unit.
The negative control gave the expected baseline value and variability, in agreement with the guideline indications. According to the method, the mean value is considered the baseline value of the experiment and thus represents 100% of cell viability.
The positive control caused the expected cell death (2.9% of cell viability when compared to the negative control) and variability (CV% equal to 12.2).
Therefore, the assay was regarded as valid.
The test item induced cell death in the three replicates with a mean cell viability of 3.8% when compared to the negative control. Intra-replicate variability was higher than expected. This may be due to the nature of the substance (cream) and thus, to the fact that residues might be present on the surface even with a higher number of washings after treatment. However, since the behavior of the three replicates indicated that viability was well below the cut-off value of 50%, the test item results were accepted as valid.
According to the established criteria (cell viability less than 50%), the test item is considered to have irritant effect on the skin under the reported experimental conditions.
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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