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EC number: 221-831-7 | CAS number: 3248-91-7
- 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:
- weight of evidence
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- data from handbook or collection of data
- Justification for type of information:
- Data is from Peer reviewed journals
Data source
Reference
- Reference Type:
- publication
- Title:
- Use of HPLC/UPLC-spectrophotometry for detection of formazan in in vitro Reconstructed human Tissue (RhT)-based test methods employing the MTT-reduction assay to expand their applicability to strongly coloured test chemicals
- Author:
- N. Alépée, J. Barroso , A. De Smedt, B. De Wever, J. Hibatallah, M. Klaric, K.R. Mewes,M. Millet, U. Pfannenbecker, M. Tailhardat, M. Templier, P. McNamee
- Year:
- 2 015
- Bibliographic source:
- Toxicology in Vitro, 29 (2015), 741–761
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Principles of method if other than guideline:
- EpiSkin™ in vitro test method was performed in 3 different labs to assess the skin irritation potential of 4-[(4-amino-m-tolyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-o-toluidine monohydrochloride
- GLP compliance:
- not specified
Test material
- Reference substance name:
- 4-[(4-amino-m-tolyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-o-toluidine monohydrochloride
- EC Number:
- 221-831-7
- EC Name:
- 4-[(4-amino-m-tolyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-o-toluidine monohydrochloride
- Cas Number:
- 3248-91-7
- Molecular formula:
- C22H24ClN3
- IUPAC Name:
- 4-[(4-amino-m-tolyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-o-toluidine monohydrochloride
- Details on test material:
- -[(4-amino-m-tolyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-o-toluidine monohydrochloride/ 3248-91-7
/ 221-831-7
Commen name-Basic Violet 2
mol.formula:C22H23N3 .HCl
smiles:C(\c1cc(c(N)cc1)C)(c1cc(c(N)cc1)C)=C1\C=C(C(=[NH+])C=C1)C.[ClH-]
InChI:1S/C22H23N3.ClH/c1-13-10-16(4-7-19(13)23)22(17-5-8-20(24)14(2)11-17)18-6-9-21(25)15(3)12-18;/h4-12,23H,24-25H2,1-3H3;1H
Physical state: Solid Green crystalline powder
molecular weight:365.906 g/mol
Constituent 1
- Specific details on test material used for the study:
- - Name of test material: C.I. Basic Violet 2 / (4-[(4-amino-m-tolyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-o-toluidine monohydrochloride)
- IUPAC name: 4-[(4-amino-3-methylphenyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-2-methylaniline hydrochloride
- Molecular formula: C22H24ClN3
- Molecular weight: 365.906 g/mole
- Smiles :C(\c1cc(c(N)cc1)C)(c1cc(c(N)cc1)C)=C1\C=C(C(=[NH+])C=C1)C.[ClH-]
- Inchl: 1S/C22H23N3.ClH/c1-13-10-16(4-7-19(13)23)22(17-5-8-20(24)14(2)11-17)18-6-9-21(25)15(3)12-18;/h4-12,23H,24-25H2,1-3H3;1H
- Substance type: Organic
- Physical state: Solid Green crystalline powder
In vitro test system
- Test system:
- other: Reconstructed human epidermis test method
- Source species:
- human
- Cell type:
- non-transformed keratinocytes
- Cell source:
- other: Not specified
- Source strain:
- other: human skin
- Details on animal used as source of test system:
- RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiSkin™ in vitro test method
TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: -20 deg C
- Wavelength and band pass (if applicable) used for quantifying MTT formazan, and linearity range of measuring device (e.g., spectrophotometer); - 570 nm
- MTT concentration: as per OECD Guidelines - Justification for test system used:
- Reconstructed Human tissue(Rht) most recently adapted on july 26th 2013
- Vehicle:
- water
- Details on test system:
- RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiSkin™ in vitro test method
TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: -20 deg C
MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: as per OECD Guidelines - Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- yes, concurrent MTT non-specific colour control
- Amount/concentration applied:
- Not specified
- Duration of treatment / exposure:
- according to the guidelines
- Duration of post-treatment incubation (if applicable):
- No data.
- Number of replicates:
- No data
Results and discussion
In vitro
Resultsopen allclose all
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 1
- Value:
- 114.2
- Vehicle controls validity:
- not specified
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Remarks:
- when applied as solid
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 2
- Value:
- 119.1
- Vehicle controls validity:
- not specified
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Remarks:
- when applied as solid
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 3
- Value:
- 2.5
- Vehicle controls validity:
- not specified
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Remarks:
- when applied as solid
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 1
- Value:
- 108.3
- Vehicle controls validity:
- not specified
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Remarks:
- when applied as a 1% (w/v) aqueous solution
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 2
- Value:
- 107.2
- Vehicle controls validity:
- not specified
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Remarks:
- when applied as a 1% (w/v) aqueous solution
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 3
- Value:
- 109.8
- Vehicle controls validity:
- not specified
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Remarks:
- when applied as a 1% (w/v) aqueous solution
- Other effects / acceptance of results:
- Acceptance criteria based on positive and negative controls in accordance with test guidelines
Any other information on results incl. tables
Table 3:EpiSkin™% tissue viability quantification from formazan tissue extracts by OD (L’Oréal R&I only) and HPLC/UPLC-spectrophotometry (3 laboratories) for the test chemicals
Name of the test chemical |
In vivo Classification |
Pre checks |
EpiSkin™ (skin irritation) Tissue viability (%) |
In vitro skin irritation classification |
OD-HPLC/UPLC spectrophotometry viability concordant Y/N |
||||||||
MTT reducer (Y/N)
|
Colour interference (Y/N) |
||||||||||||
OD analysis L’Oréal R&I |
HPLCanalysis L’Oréal R&I |
HPLCanalysis L’Oréal R&I |
UPLC analysis VITO |
HPLC/UPLC spectrophotometry SD (3 Labs)* |
Difference (OD-HPLC) L’Oréal R&I |
OD |
HPLC/ UPLC spectrophotometry |
||||||
Benzenamine, 4-((4-amino-3-methyl-phenyl)(4-imino-3-methyl-2,5-cyclohexadiene-1-ylidene)methyl-2-methyl HCl |
NC |
N |
Y |
159.0b(228.6) |
114.2 (114.2) |
119.1 (119.1) |
2.5 |
44.8 |
119.1 (119.1) |
Not compatible |
NC |
NA |
|
Benzenamine, 4-((4-amino-3-methyl-phenyl)(4-imino-3-methyl-2,5-cyclohexadiene-1-ylidene)methyl-2-methyl HCl solution 1% (w/v) aqueous |
NC |
N |
Y |
99.6b(233.4) |
108.3 (108.3) |
107.2 (107.2) |
109.8 (109.8) |
1.3 |
8.7 |
Not compatible |
NC |
NA |
Where
Uncorrected viability values in parentheses.
Not compatible: Using the OD measurement (% NSC and/or % NSMTT≥50%).
Y: Yes.
N: No.
NA: Not applicable.
NK: Not known.
NT: Not tested.
ND: Could not be determined.
Cat 1: Skin Corrosive.
NC: Not Corrosive.
NC: Not Classified.
%NSMTT≥50%
a- SD between three replicate tissues ≥18%
b - NSC≥50%.
c - NSMTTP50%.
** OD-HPLC/UPLC-spectrophotometry > 20%.
*** NT based on a decision by the participating laboratory not to proceed to further testing of this test chemical.
**** Difference between 2 laboratories.
Applicant's summary and conclusion
- Interpretation of results:
- other: Not irritating
- Conclusions:
- The mean % tissue viability of C.I. Basic Violet 2 when evaluated as a solid in 3 different laboratories were 114.2, 119.1, 2.5 respectively and 108.3, 107.2, 109.8 respectively when evaluated as a 1% (w/v) aqueous solution. The results obtained from the test indicated a very strong possibility of Basic Violet 2 being not irritating to skin.
- Executive summary:
EpiSkin™ in vitro test method was performed in 3 different labs to assess the skin irritation potential of4-[(4-amino-m-tolyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-o-toluidine monohydrochloride. Since, 4-[(4-amino-m-tolyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-o-toluidine monohydrochloride[C.I. Basic Violet 2] exhibited extreme color interference when tested undiluted, hence it was also evaluated as a 1% (w/v) aqueous dilution. This was done to determine how strong the colour interference °remained with dilution and the relevance of the two endpoint detection systems (OD and HPLC/UPLC-spectrophotometry).
EpiSkin™ in vitro test method was performed using the SOPs in accordance with the relevant OECD test guidelines i.e. Updated OECD TG439.Both positive and negative controls were run in parallel with the test substances. Acceptance criteria based on positive and negative controls were fulfilled as mentioned in the SOPs. The evaluation of direct MTT reduction involved the incubation of the test chemical with MTT solution. If the solution turned blue, killed tissue adapted controls incubated with MTT to determine non-specific MTT reduction (% NSMTT).Since the test chemical was both colored and MTT reducer, living tissue adapted controls to define non-specific colour (% NSC) and killed tissue adapted controls to determine non-specific MTT reduction (% NSMTT) were deemed necessary. Also a third set of adapted controls using killed tissues incubated with medium instead of MTT were used to define chemical binding to the killed tissue (% NSCkilled). In this case, the final corrected % tissue viability for the test chemical was obtained by subtracting % NSC and % NSMTT and adding % NSCkilled to the % tissue viability obtained with the chemical treated living tissues incubated with MTT.
The study was performed by L’Oreal R&I.The resulting formazan tissue extracts were analysed by photometry (OD) in accordance with the SOP for each test method after testing. After shipment of the formazan tissue extracts stored at -20°C, measurement of the formazan by HPLC/UPLC-spectrophotometry in the three different laboratories (L’Oréal R&I, Pierre Fabre Laboratories and VITO) was performed within one week of each other.Furthermore, photometry (OD) was performed in parallel with the HPLC/UPLC-spectrophotometric analysis of the -20°C stored samples by the laboratory that conducted the in vitro biological test method. Correlation analyses for both formazan detection methods were performed.
The mean % tissue viability of C.I. Basic Violet 2 when evaluated as a solid in 3 different laboratories were 114.2, 119.1, 2.5 respectively and 108.3, 107.2, 109.8 respectively when evaluated as a 1% (w/v) aqueous solution. The results obtained from the test indicated a very strong possibility of Basic Violet 2 being not irritating to skin.
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