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EC number: 254-991-1 | CAS number: 40594-65-8
- 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 sensitisation
Administrative data
- Endpoint:
- skin sensitisation: in vitro
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- test procedure in accordance with national standard methods with acceptable restrictions
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Report date:
- 2014
Materials and methods
Test guideline
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- other: OECD 442E (In Vitro Skin Sensitisation: human Cell Line Activation Test (h-CLAT))
- Principles of method if other than guideline:
- The evaluation of pro-sensitising activity of the substance on human monocytes through FACS analysis was performed.
- GLP compliance:
- no
- Remarks:
- not-GLP study but considered as reliable based on the reporting of the study
- Type of study:
- activation of dendritic cells
Test material
- Reference substance name:
- 5-methyl-2-(isopropyl)cyclohexyl nicotinate
- EC Number:
- 254-991-1
- EC Name:
- 5-methyl-2-(isopropyl)cyclohexyl nicotinate
- Cas Number:
- 40594-65-8
- Molecular formula:
- C16H23NO2
- IUPAC Name:
- 5-methyl-2-(isopropyl)cyclohexyl nicotinate
Constituent 1
In vitro test system
- Details on the study design:
- CELL MODEL: monocyte-like human line called THP-1. Cells are cultured in RPMI containing 10 % FCS and 2 mM glutamine.
PRELIMINARY CYTOTOXICITY STUDY: after exposure of the cells to the test material, the cells are washed with PBS and exposed to the MTT-medium at 37°C. At the end of the incubation period, the MTT-medium is removed and the cells receive the MTT solubilization solution. The plate is shaken on a rotatory plate shaker for 20-30 minutes, ensuring that all the crystals have dissolved from the cells and have formed a homogeneous solution. The absorbance is measured as described with background elimination. The results are expressed in terms of viability: % viability = (OD treated cultures*100)/OD untreated control cultures.
SENSITISATION ASSAY:
-Sample preparation: the sample was dissolved in ethanol and then diluted in the cell medium at the two different dilutions in order to obtain the desired final concentrations in contact with the THP-1cells in vitro.
- Test concentrations: 3 μg/ml and 0.6 μg/ml.
- Exposure period: 48 h.
- Exposure conditions: at 37 °C with 5 % CO2.
- Staining: after the incubation with the test substance and the controls, cells were collected, checked under the microscope for their vitality by staining with Trypan Blue dye and counted in a cell counter chamber.
- Measurement of markers expression: after staining the cells were washed in PBS and then marked with a fluoresceinated anti-B7.1 or B7.2 antibody. After washing, to eliminate the excess antibody, the MFI (Mean Fluorescence Intensity) linked to the cells was evaluated by means of a flow cytofluorimeter (FACS, Fluorescence Activated Cell Sorter, Becton Dickinson, Mountain View, CA). This value is proportional to the expression of costimulatory molecules. The MFI of the non-treated THP-1 cells and of cells after reaction with a monoclonal isotype-matched antibody was used as an internal control (basal fluorescence). MFI is the geometric average of the fluorescence intensity of the cells treated with the fluoresceinated antibody and it is proportional to the number of stained molecules per cell.
- Positive control: nickel sulphate.
- Negative control: cell medium, non treated cells.
Results and discussion
- Positive control results:
- Test on nickel sulphate, a known sensitising substance, is characterised by a) high increase of both the markers; b) direct correlation between concentration and intensity of the response; c) relevant effects even at very low doses. The tested dose of 4 mg/ml of Nickel Sulphate (NiSO4.6H2O) corresponds to approx. 1 ppm of Nickel, concentration that is around the minimal sensitising threshold in already sensitised individuals with irritated skin (1,2). A low dose of nickel sulphate can cause a detectable increase of CD80 respect to untreated cell. The concentration that is able to cause an allergic reaction in most of the sensitive subjects in intact skin is over the 100 ppm of Nickel Sulphate.
1. Gawkrodger DJ, Nickel dermatitis: how much Nickel is safe? Contact Dermatitis 1996; 35: 267-271.
2. Tanojo H, Hostynek JJ, Mountford HS, Maibach HI. In vitro permeation of nickel salts through human stratum corneum. Acta Derm Venereol Suppl, 2001; 212: 19-23.
In vitro / in chemico
Resultsopen allclose all
- Parameter:
- other: MFI (CD80)
- Value:
- -0.42
- Vehicle controls validity:
- not specified
- Negative controls validity:
- not specified
- Positive controls validity:
- valid
- Remarks on result:
- other: MFI values obtained for test conc. 3 and 0.6 μg/ml/.
- Parameter:
- other: MFI (CD86)
- Value:
- -0.82
- Vehicle controls validity:
- not specified
- Negative controls validity:
- not specified
- Positive controls validity:
- valid
- Remarks on result:
- other: MFI values obtained for test conc. 3 and 0.6 μg/ml/.
- Other effects / acceptance of results:
- The sample did not show cytotoxic effects on the cells used for the test (IC50 = 15 μg/ml)
No signal related to apoptosis on the treated cells was observed.
The sample did not show any increase in the expression of the investigated markers.
Any other information on results incl. tables
The results expressed as co-stimulatory molecules expression after the exposure to the substance for the positive control substance and test substance are presented in the table below.
Table
Sample | CD80 (MFI) | CD86 (MFI) |
Nickel sulphate 20 μg/ml |
11.35 | 57.66 |
Nickel sulphate 10 μg/ml |
9.63 | 24.88 |
Nickel sulphate 4 μg/ml |
1.43 | 11.39 |
Test material 3 μg/ml |
-0.42 | -0.82 |
Test material 0.6 μg/ml |
-0.33 | -0.8 |
Applicant's summary and conclusion
- Interpretation of results:
- other: not considered to be a skin sensitiser according to the in vitro study results
- Conclusions:
- The substance did not show any increase in the expression of the investigated markers. It is not considered to have a sensitising potential.
- Executive summary:
The sensitising potential of the substance was evaluated in-vitro by quantifying the changes of cell surface marker expression (CD80, CD86) on a human monocytic leukemia cell line, THP-1 cells, following 48 hours exposure to the two concentrations of the test substance. The changes of surface marker expression were measured by a flow cytofluorimeter following cell staining with fluoresceinated antibodies. A preliminary cytotoxicity measurement (MTT assay) was also conducted to determine the non-cytotoxic concetrations to be used in the sensitisation assay. The Mean Fluorescence Intensity (MFI) of surface markers for the test material and positive control (nickel sulphate) compared to negative control were calculated.
The results of nickel sulphate is characterised by a) high increase of both the markers; b) direct correlation between concentration and intensity of the response; c) relevant effects even at very low doses (4 μg/ml). The sample did not show any increase in the expression of the markers CD80 and CD86.
Comparing the behaviour of the positive substance and the test material it is deemed that the substance does not have sensitising potential.
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