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EC number: 273-453-7 | CAS number: 68966-86-9
- 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:
- 15 February 2016 - 22 February 2016
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- According to Annex VII of the REACH Regulation, if new test data are required these must be derived from in vitro methods only.
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 016
- Report date:
- 2016
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:
- Based on a "Statement on the Scientific Validity of In Vitro Tests for Skin Irritation" (November 2008)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: UN GHS
- Version / remarks:
- 3rd revision (2009)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
Test material
- Reference substance name:
- Methyl 4(or 1)-isopropyl-1(or 4)-methylbicyclo[2.2.2]oct-5-ene-2-carboxylate
- EC Number:
- 273-453-7
- EC Name:
- Methyl 4(or 1)-isopropyl-1(or 4)-methylbicyclo[2.2.2]oct-5-ene-2-carboxylate
- Cas Number:
- 68966-86-9
- Molecular formula:
- C14H22O2
- IUPAC Name:
- Methyl 4(or 1)-isopropyl-1(or 4)-methylbicyclo[2.2.2]oct-5-ene-2-carboxylate
Constituent 1
In vitro test system
- Test system:
- human skin model
- Source species:
- human
- Cell type:
- non-transformed keratinocytes
- Cell source:
- other:
- Source strain:
- other:
- Details on animal used as source of test system:
- SOURCE ANIMAL
- Source: Human donation
- Tissue: normal epidermal keratinocytes - Justification for test system used:
- Elicited via a disturbance of the desquamation process and an inflammatory response (i.e. papules, vesicles, bullae and oedema), skin irritation requires penetration of the stratum corneum and elicitation of a biological response. Skin irritation is defined in Section 3.2.1.1 of Annex I to the CLP regulation as “...the production of reversible damage of the skin following the application of a test substance for up to 4 hours”. The EpiDerm™ human skin model (OECD 439) is an accepted in vitro test method to detect skin corrosion/irritation (Category 1 or 2) and/or the absence of effects (not classified under CLP).
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: Epi-200 SIT Kit
- Tissue batch number(s): 23314
- Production date: Not reported
- Shipping date: Not reported
- Delivery date: 16 February 2016
- Date of initiation of testing: Pre-incubation phase started 16 February 2016
TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37 ± 1.5 ºC
- Temperature of post-treatment incubation (if applicable): 37 ± 1 ºC
REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: 300 μL, one washing step
- Observable damage in the tissue due to washing: None reported
- Modifications to validated SOP: None
MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: 3 hours
- Spectrophotometer: Not applicable, microplate reader used
- Wavelength: 570 ± 1 nm
- Filter: Not reported
- Filter bandwidth: 570 ± 1 nm
- Linear OD range of spectrophotometer: Not reported
FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: Positive control; 4.77 %
- Barrier function: Not reported
- Morphology: Not reported
- Contamination: None reported
- Reproducibility: Relative standard deviation for mean viability; 14.79 % postive control. Relative standard deviation for mean absorption; 16.6 % postive control, 8.55 % negative control.
NUMBER OF REPLICATE TISSUES: 3 per control and test item treatment
NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 3
PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be corrosive to skin if the viability after 43.5 hours exposure is less than 50% - Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- Amount/concentration applied:
- TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 30 μL
- Concentration (if solution): Undiluted test item
NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 μL
- Concentration (if solution): Not reported
POSITIVE CONTROL
- Amount(s) applied (volume or weight): 30 μL
- Concentration (if solution): 5 % SLS in deioinsied water; concentration not reported - Duration of treatment / exposure:
- 43.5 hours
- Duration of post-treatment incubation (if applicable):
- 67.5 hours
- Number of replicates:
- 3 per test item, negative or positive control
Results and discussion
In vitro
Resultsopen allclose all
- Irritation / corrosion parameter:
- other: mean tissue absorbance
- Remarks:
- Of three wells after blank corrction
- Run / experiment:
- 1
- Value:
- 1.615
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- other: Mean tissue absorbance
- Remarks:
- Of three wells after blank
- Run / experiment:
- 2
- Value:
- 1.338
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- other: Mean tissue absorbance
- Remarks:
- Of three wells after blank correction
- Run / experiment:
- 3
- Value:
- 1.495
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- other: Mean tissue absorbance
- Remarks:
- Mean of three tissue after blank correction
- Run / experiment:
- Mean
- Value:
- 1.483
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- other: Relative absorbance (%)
- Remarks:
- compared to blank
- Run / experiment:
- TIssue
- Value:
- 103.4
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- other: Relative absorbance (%)
- Remarks:
- compared to blank
- Run / experiment:
- Tissue 2
- Value:
- 85.6
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- other: Relative absorbance (%)
- Remarks:
- compared to blank
- Run / experiment:
- Tissue 3
- Value:
- 95.7
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- other: Mean relative abosrbance
- Remarks:
- % of negative control
- Run / experiment:
- Mean
- Value:
- 94.9
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Other effects / acceptance of results:
- - OTHER EFFECTS:
- Visible damage on test system: None reported
- Direct-MTT reduction: Did not show blue colour after 1 hour incubation
- Colour interference with MTT: No colour change
DEMONSTRATION OF TECHNICAL PROFICIENCY: Not reported
ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes, within the required range of OD ≥ 0.8 and ≤ 2.8
- Acceptance criteria met for positive control: Yes, induced a decrease in absorbance of 5.1 % compared to the negative control
- Acceptance criteria met for variability between replicate measurements: Not reported
- Range of historical values if different from the ones specified in the test guideline:Not applicable
Any other information on results incl. tables
Results of treatment with test item and the controls
Dose Group | Exposure Interval | Tissue No. | Absorbance 570 nm (Well 1) | Absorbance 570 nm (Well 2) | Absorbance 570 nm (Well 3) | Mean Absorbance of 3 Wells* | Mean Absorbance of three wells blank corrected | Mean Absorbance of three tissues after blank correction | Rel. Absorbance (%) 1, 2 + 3** | Relative Standard Deviation (%) | Mean. Rel. Absorbance (% of negative control)*** |
Blank | 0.037 | 0.038 | 0.038 | 0.038 | 0.000 | ||||||
Negative Control | 60 mins | 1 | 1.540 | 1.548 | 1.508 | 1.532 | 1.495 | 1.563 | 95.6 | 4.1 | 100.0 |
2 | 1.644 | 1.591 | 1.593 | 1.609 | 1.572 | 100.6 | |||||
3 | 1.717 | 1.631 | 1.630 | 1.659 | 1.622 | 103.8 | |||||
Positive Control | 60 mins | 1 | 0.117 | 0.120 | 0.118 | 0.118 | 0.081 | 0.080 | 5.2 | 4.4 | 5.1 |
2 | 0.118 | 0.107 | 0.116 | 0.113 | 0.076 | 4.8 | |||||
3 | 0.104 | 0.132 | 0.124 | 0.120 | 0.083 | 5.3 | |||||
Test item | 60 mins | 1 | 1.672 | 1.634 | 1.652 | 1.653 | 1.615 | 1.483 | 103.4 | 9.4 | 94.9 |
2 | 1.384 | 1.362 | 1.380 | 1.375 | 1.338 | 85.6 | |||||
3 | 1.530 | 1.528 | 1.540 | 1.532 | 1.495 | 95.7 |
* Mean of three replicate wells after blank correction
** relative absorbance per tissue (rounded values): 100*(absorbancetissue)/ (mean absorbancenegative control)
***relative absorbance per treatment group (rounded values): 100*(mean absorbancetest item/positive control)/ (mean absorbancenegative control)
Applicant's summary and conclusion
- Interpretation of results:
- GHS criteria not met
- Conclusions:
- The test item was non-cytotoxic in an experiment employing an artificial three-dimensional model of human skin. Conducted according to OECD 439 (EU B.46) and GLP, the in vitro study is considered reliable without restriction (Klimisch 1). The test item should not be classified as an irritant (UN GHS no category).
- Executive summary:
Skin irritation of the test item was evaluated with the EpiDerm Reconstructed Human Epidermis Model. Cell viability of the multi-layered tissue culture of highly differentiated epidermal keratinocytes topically exposed to the test substance was evaluated using the MTT assay, which measures the conversion of 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl-tetrazoliumbromide (MTT) into a blue formazan salt. Undiluted test item was applied to the EpiDerm tissue for 60 minutes, alongside a negative and positive control. The mean relative absorbance value of the test item, corresponding to the cell viability did not significantly decrease (94.9%; threshold for irritancy: ≤ 50%), consequently the test item was not irritant to skin. The test item passed the MTT- and the Colour Interference pre-tests. Conducted according to OECD Test Guideline 439 and GLP, the study is considered to be reliable without restriction (Klimisch 1).
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