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EC number: 248-702-8 | CAS number: 27870-92-4
- 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
Endpoint summary
Administrative data
Description of key information
Skin irritation:
The dermal irritation potential of test article 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined according to the OECD 439 test guideline followed for this study. The Mean % tissue viability compared to negative control (n=3) of the test subst ance 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione was determined to be 103.5 %. Thus, 2-octadecyl-1H-thioxantheno [2,1, 9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was considered to be not irritating to the skin of humans.
Eye irritation:
The ocular irritation potential of test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined according to the OECD 492 test guideline followed for this study. The mean % tissue viability of test substance 2-octadecyl -1H-thioxantheno [2,1,9- def]isoquinoline-1,3(2H)-dione was determined to be 91.5%. Thus, substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) is considered to be "not irritating" to the human eye.
Key value for chemical safety assessment
Skin irritation / corrosion
Link to relevant study records
- Endpoint:
- skin irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- May 05, 2017 to July 17, 2017
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- Data is from experimental study report
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Principles of method if other than guideline:
- The purpose of this study was to assess potential for the test articles to be dermal irritants. The dermal irritation potential of test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) may be predicted by measurement of their cytotoxic effect, as reflected in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, in the MatTek EpiDerm™ model (MatTek Corp., Ashland, MA).
- GLP compliance:
- yes
- Specific details on test material used for the study:
- SOURCE OF TEST MATERIAL
- Test material identity: 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (Neeliglow. Solvent Yellow 98)
- Source: Neelikon Food Dyes & Chemicals Ltd.
- Lot No.of test material: FG/16-17/0289
- Date of manufacture: 27/04/2016
- Expiration date of the lot/batch: April 27, 2026
- Purity: Not provided
- Melting point: 108 degC (minimum)
- Melting point: 110 degC (maximum)
- Absorbance for 5 ppm solution in Dimethyl Formamide: 0.189 A°
- Wavelength maxima in DMF: 457 nm
RADIOLABELLING INFORMATION (Not applicable)
- Radiochemical purity: N/A
- Specific activity: N/A
- Locations of the label: N/A
- Expiration date of radiochemical substance: N/A
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature
- Stability under test conditions: No data available
- Solubility and stability of the test substance in the solvent/vehicle: No data available
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: No data available
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Test articles is tested as provided (neat).
- Preliminary purification step (if any): No data available
- Final dilution of a dissolved solid, stock liquid or gel: No data available
- Final preparation of a solid: No data available
FORM AS APPLIED IN THE TEST (if different from that of starting material): Solid
OTHER SPECIFICS: No data available - Test system:
- human skin model
- Source species:
- human
- Cell type:
- non-transformed keratinocytes
- Cell source:
- other: EpiDerm™ 3-dimensional human tissues used in this study
- Source strain:
- other: Not applicable
- Details on animal used as source of test system:
- - Description of the cell system used:
The normal human-derived keratinocytes were cultured at the air-liquid interface in a chemically defined medium on a permeable polycarbonate insert (surface 0.5 cm2). They were cultured in chemically defined serum free medium to form a multi-layered epithelium similar to that found in native epidermis. Each lot of tissues was Quality Assured by MatTek according to specific QC standards including: histology, tissue viability (MTT mean optical density), reproducibility (SD) and tissue thickness.
- Test System Identification
All of the EpiDerm™ 3-dimensional human tissues used in this study were identified by the date of arrival and the lot number. Certificate of Analysis for the tissues are included in this report. Tissue plates were appropriately labeled with study information. - Justification for test system used:
- The 3-Dimensional Human Dermal Epithelial Model (EpiDerm™, MatTek, Ashland, MA) is made up of normal human keratinocytes in serum free medium. The cells form an epithelial tissue that consists of organized basal, spinous, granular, and cornified layers analogous to those found in vivo. The EpiDerm™ model also contains epidermis-specific differentiation markers such as pro-filaggrin, the K1/K10 cytokeratin pair, involucrin, and type I epidermal transglutaminase, as well as keratohyalin granules, tonofilament bundles, desmosomes, and a multi-layered stratum corneum containing intercellular lamellar lipid layers arranged in patterns characteristic of in vivo epidermis. Each lot of tissues was Quality Assured by MatTek, Inc. according to specific QC standards including: histology (cell layers), tissue viability (MTT mean optical density) and reproducibility (SD). Tissue plates were appropriately labeled with study information. Bias was not a factor in this test system.
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- The tissues were exposed to the test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione neat (undiluted) on June 28, 2017 (Run 1 of 1). EpiDerm™ tissues were purchased from MatTek. Quality control of the tissues was performed by MatTek and the Certificate of Analysis (CoA) for the tissues is provided and is kept in the study binder. Tissues were exposed for approximately 1 hour, with 35 minutes in an approximately 37°C, 5% CO2 humidified incubator and the remaining 25 minutes at room temperature. Following the exposure time, the tissues were rinsed and placed in fresh media for approximately 24 hours. The media was then changed again and the tissues were incubated in fresh media for another ~18 hours for a total of approximately 42 hour post-exposure recovery period. The tissue viability was then assessed by MTT assay. The tissue CoA was used instead of verification of barrier properties of the tissue.
MTT and Color Pre-tests
Pretesting for MTT auto-reduction and coloring was not performed for this study but was based on the results obtained from another study (CYP1690_R1b).
MTT Assay
Following the rinsing period, the MTT assay was performed by transferring the tissues to 24-well plates containing 300 µL MTT medium (1.0 mg/mL). After 2 hours, 57 minute and 25 second MTT incubation at approximately 37°C, approximately 5% CO2 in a humidified incubator, the blue formazan salt was extracted by submerging tissues in 2 mL isopropanol in a 24-well plate. The extraction time was approximately 2 hours 04 minutes and 11 seconds with gentle shaking. The optical density of the extracted formazan (200 µL/well of a 96-well plate) was determined using a Synergy H4 spectrophotometer at 570 nm. Relative cell viability is calculated for each tissue as % of the mean negative control tissues.
Evaluation of Test Article in the Cell Models:
1. Cell system:
Upon receipt, the MatTek EpiDerm™ tissue cultures were placed in 0.9 mL of fresh Maintenance medium (in a 6-well plate). The culture inserts are incubated for ~one hour. The tissues were then transferred to 6-well plates containing 0.9 mL fresh Maintenance medium and they were incubated overnight at ~37°C, 5% CO2 in a humidified incubator.
2. Control and Test Article Exposures:
On the day of dosing, the tissues are then removed from the incubator and the controls and the test articles are applied topically to tissues by pipette. Tissues were exposed to controls and the test articles for one hour, with ~35 minutes in a 37°C, 5% CO2 humidified incubator and the remaining 25 minutes at room temperature.
a) Controls
30 µL of negative control DPBS, positive control 5% SDS was applied topically to the tissue and gently spread by placing a nylon mesh on the apical surface of each tissue, if necessary.
b)Test Article
For 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione, the tissues were moistened with 25 μL of ultrapure water to improve contact of the tissue surface with the test article. Approximately 25 mg of 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione was evenly applied to the apical surface of each tissue (n=3). All the tissues were placed into the ~37°C incubator with 5% CO2. The exposure times were approximately 1 hour, with ~35 minutes exposure in the incubator and ~25 minutes at room temperature.
3.Post-exposure treatment
After the 1 hour exposure, the tissues were rinsed 20 to 25 times with 1 mL of DPBS. The apical surface was gently blotted with a cotton swab. The tissues were placed in 0.9 mL of fresh Maintenance medium (6-well plate) for either 25 hours, 38 minutes and 23 seconds or for 24 hours, 10 minutes and 09 seconds (as there were numerous tissues, they had to be broken down into 2 sets to complete dosing in a timely manner). After this initial ~24 hour incubation, the tissues were placed in 6-well plates containing 0.9 mL fresh Maintenance medium and incubated for another 17 hours, 03 minutes and 34 seconds prior to performing the MTT assay, for a total of an approximately 42 hour post-exposure incubation.
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: The EpiDerm™ 3 dimensional human tissue model
- Tissue Lot number(s): 26459
- Date of initiation of testing: 6/08/2017
TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37°C
- Temperature of post-treatment incubation (if applicable): 37°C
REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: Twice
MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 300 µL MTT medium (1.0 mg/mL).
- Incubation time: After 2 hours, 57 minute and 25 second MTT incubation
- Spectrophotometer: Synergy H4 spectrophotometer
- Wavelength: 570 nm
- Filter: No data
- Filter bandwidth: No data
- Linear OD range of spectrophotometer: No data
NUMBER OF REPLICATE TISSUES: 3
CALCULATIONS and STATISTICAL METHODS
All data were background subtracted before analysis. MTT data are presented as % viable compared to negative control. Data were generated as follows:
MTT Assay
Blanks:
· The optical density (OD) mean from all replicates for each plate (ODblank).
Negative Controls (NC):
· The blank corrected value was calculated: ODNC= ODNCraw– ODblank.
· The OD mean per NC tissue was calculated.
· The mean OD for all tissues corresponds to 100% viability.
· The mean, standard deviation (SD), standard error of the mean (SEM) and the percent coefficient of variation (% CV) was calculated.
Positive Control (PC):
· Calculate the blank corrected value: ODPC= ODPCraw– ODblank.
· The OD mean per PC tissue was calculated.
· The viability per tissue was calculated: %PC = [ODPC/ mean ODNC] x 100.
· The mean viability for all tissues was calculated: Mean PC = Σ %PC / number of tissues.
· The standard deviation (SD), standard error of the mean (SEM) and the percent coefficient of variation (% CV) was calculated.
Tested compound :
· Calculate the blank corrected value ODTT= ODTTraw– ODblank.
· The OD mean per tissue was calculated.
· The viability per tissue was calculated: %TT = [ODTT/ mean ODNC] x 100.
· The mean viability for all tissues was calculated: Mean TT = Σ %TT / number of tissues.
· The standard deviation (SD), standard error of the mean (SEM) and the percent coefficient of variation (% CV) was calculated.
Data Correction Procedure for MTT Interfering Compounds (if applicable)
True viability = Viability of treated tissue – Interference from test article = ODtvt– ODktwhere ODkt= (mean ODtkt– mean ODukt).
ODtvt= optical density of treated viable tissue
ODkt= optical density of killed tissues
ODtkt= optical density of treated killed tissue
ODukt= optical density of untreated killed tissue (NC treated tissue)
Data Correction Procedure for Colored Compounds (if applicable)
True viability = Viability of treated tissue incubated in MTT media – Viability of treated tissue incubated in media without MTT = ODtvt– ODvt.
ODtvt= optical density of treated viable tissue incubated in MTT media
ODvt= optical density of viable tissues incubated in media alone
- Evaluation of data
The results of the assay was evaluated and compared to negative control.
Table: Criteria for in vitro Interpretation:
In VitroResults In VivoPrediction
Mean tissue viability ≤50% Irritant (I), R38
Mean tissue viability >50% Non-irritant (NI)
- Assay quality controls
- Negative Controls (NC)
The Dulbecco’s phosphate buffered saline (DPBS) was used as a NC. The assay passed all acceptance criteria if the ODs of the negative control exposed tissues were between ≥0.8 and ≤2.8.
- Positive Controls (PC)
5% solution of sodium dodecyl sulfate was used as a PC. The assay is meeting the acceptance criteria if the viability of the PC is ≤20% of the negative control.
- Standard Deviation (SD)
The standard deviation (SD) calculated from individual percent tissue viabilities of the test article exposed replicates was ≤18. - Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- Amount/concentration applied:
- TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 mg
- Concentration (if solution): neat (undiluted)
VEHICLE (Not used)
- Amount(s) applied (volume or weight with unit): none
- Concentration (if solution): none
- Lot/batch no. (if required): none
- Purity: none
NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 µL
- Concentration (if solution): neat
POSITIVE CONTROL
- Amount(s) applied (volume or weight): 30 µL
- Concentration (if solution): 5% solution of sodium dodecyl sulfate - Duration of treatment / exposure:
- The exposure times were approximately 1 hour, with ~35 minutes exposure in the incubator and ~25 minutes at room temperature.
- Duration of post-treatment incubation (if applicable):
- For a total of an approximately 42 hour post-exposure incubation.
- Number of replicates:
- 3 tissues were used for test compound and control.
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- Run 1
- Value:
- 103.5
- Vehicle controls validity:
- not specified
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Other effects / acceptance of results:
- The MTT data show the assay quality controls were met, as the OD of the negative control tissues was between 1.195 and 1.430. Also, the positive control, 5% sodium dodecyl sulfate (SDS), reduced tissue viability to 4.5% of negative control and the standard deviation (SD) calculated from individual percent tissue viabilities of the test article exposed replicates was 2.9 passing the acceptance criteria.
- Interpretation of results:
- other: not irritating
- Conclusions:
- The dermal irritation potential of test article 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined according to the OECD 439 test guideline followed for this study. The Mean % tissue viability compared to negative control (n=3) of the test subst ance 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione was determined to be 103.5 %. Thus, 2-octadecyl-1H-thioxantheno [2,1, 9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was considered to be not irritating to the skin of humans.
- Executive summary:
The dermal irritation potential of test article 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined according to the OECD 439 test guideline followed for this study. The MatTek EpiDerm™ model was used to assess the potential dermal irritation of the test article by determining the viability of the tissues following exposure to the test article via MTT. The objective of this study was to assess the dermal irritation potential of test article. Tissues were exposed to test article and controls for ~one hour, followed by a 42 hour post-exposure recovery period. The viability of each tissue was determined by MTT assay.
The MTT data show the assay quality controls were met, as the OD of the negative control tissues was between 1.195 and 1.430. Also, the positive control, 5% sodium dodecyl sulfate (SDS), reduced tissue viability to 4.5% of negative control and the standard deviation (SD) calculated from individual percent tissue viabilities of the test article exposed replicates was 2.9 passing the acceptance criteria.
The Mean % tissue viability compared to negative control (n=3) of the test substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione was determined to be 103.5 %.
Hence, under the experimental test conditions it was concluded that test substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was considered to be not irritating to the human skin and being classified as “Not Classified'' as per CLP Regulation.
Reference
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (not irritating)
Eye irritation
Link to relevant study records
- Endpoint:
- eye irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- May 05, 2017 to July 12, 2017
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- Data is from experimental study report.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)
- Principles of method if other than guideline:
- The purpose of this study was to assess potential for the test article to be ocular irritants. The ocular irritation potential of a test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) may be predicted by measurement of its cytotoxic effect, as reflected in the 3-(4,5-dimethyl thiazol-2-yl)- 2,5-diphenyltetrazolium bromide (MTT) assay, in the MatTek EpiOcular™ model (MatTek Corp., Ashland, MA).
- GLP compliance:
- yes
- Specific details on test material used for the study:
- SOURCE OF TEST MATERIAL
- Test material identity: 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (Neeliglow. Solvent Yellow 98)
- Source: Neelikon Food Dyes & Chemicals Ltd.
- Lot No.of test material: FG/16-17/0289
- Date of manufacture: 27/04/2016
- Expiration date of the lot/batch: April 27, 2026
- Purity: Not provided
- Melting point: 108 degC (minimum)
- Melting point: 110 degC (maximum)
- Absorbance for 5 ppm solution in Dimethyl Formamide: 0.189 A°
- Wavelength maxima in DMF: 457 nm
RADIOLABELLING INFORMATION (Not applicable)
- Radiochemical purity: N/A
- Specific activity: N/A
- Locations of the label: N/A
- Expiration date of radiochemical substance: N/A
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature
- Stability under test conditions: Stable under normal conditions.
- Solubility and stability of the test substance in the solvent/vehicle: No data available
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: No data available
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Test articles is tested as provided (neat).
- Preliminary purification step (if any): No data available
- Final dilution of a dissolved solid, stock liquid or gel: No data available
- Final preparation of a solid: No data available
FORM AS APPLIED IN THE TEST: Powder
OTHER SPECIFICS: No data available - Species:
- human
- Strain:
- other: Not applicable
- Details on test animals or tissues and environmental conditions:
- - Description of the cell system used
The normal human-derived keratinocytes were cultured at the air-liquid interface in a chemically defined medium on a permeable polycarbonate insert (surface 0.5 cm2). They were cultured in chemically defined serum free medium to form a multi-layered epithelium similar to that found in native corneal mucosa. Each lot of tissues was Quality Assured by MatTek according to specific QC standards including: histology, tissue viability (MTT mean optical density), reproducibility (SD) and tissue thickness.
- Test System Identification
All of the EpiOcular™ 3-dimensional human tissues used in this study were identified by the date of arrival and the lot number. Certificate of Analysis for the tissues is included in this report. Tissue plates were appropriately labeled with study information. Bias was not a factor in this test system.
- Justification of the test method and considerations regarding applicability
Human Corneal Epithelia (HCE) by MatTek, Inc.:
The test article and controls were evaluated for potential ocular irritancy using the EpiOcular™ 3 dimensional human tissue model purchased from MatTek Corporation (Ashland, MA). This model consists of normal human keratinocytes cultured on a permeable synthetic membrane at the air-liquid interface in a chemically defined medium. The cells form a stratified, squamous corneal epithelium resembling the corneal mucosa of the human eye. This model has been used with several common tests of cytotoxicity including MTT and interleukin 1-alpha (IL-1α). A growing body of evidence indicates that EpiOcular™ effectively provides a non-animal means to assess potential irritancy. The EpiOcular™ model closely mimics the human corneal mucosa and thus provides an important in vitro approach in the evaluation of ocular irritancy and toxicity. Each lot of tissues was Quality Assured by MatTek, Inc. according to specific QC standards including: histology (cell layers), tissue viability (MTT mean optical density) and reproducibility (SD). - Vehicle:
- unchanged (no vehicle)
- Controls:
- other: See ''Remark" for Control Samples used in the study
- Amount / concentration applied:
- TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 50 mg
- Concentration (if solution): neat (undiluted)
VEHICLE (no vehicle)
- Amount(s) applied (volume or weight with unit): none
- Concentration (if solution): none
- Lot/batch no. (if required): none
- Purity: none
NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL
- Concentration (if solution): neat
POSITIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL
- Concentration (if solution): neat - Duration of treatment / exposure:
- Tissues were exposed for approximately ~6 hours for test article and controls, at approximately 37°C, 5% CO2 in a humidified incubator.
- Observation period (in vivo):
- Not applicable
- Duration of post- treatment incubation (in vitro):
- Following the post soak, the tissues were rinsed and incubated at approximately 37°C, 5% CO2 in a humidified incubator for a post-exposure recovery time totaling ~18 hours for test article and controls.
- Number of animals or in vitro replicates:
- 3 tissues were used for test compound and control.
- Details on study design:
- - Details of the test procedure used
The tissues were exposed to the test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione neat (undiluted). EpiOcular™ tissues were purchased from MatTek. Quality control of the tissues was performed by MatTek and the Certificate of Analysis (CoA) for the tissues is provided and is kept in the study binder. Tissues were exposed for approximately ~6 hours for test article and controls, at approximately 37°C, 5% CO2 in a humidified incubator. After the exposure, the test article was rinsed off the tissues and the tissues were soaked in media for ~25 minutes for test article and controls. Following the post soak, the tissues were rinsed and incubated at approximately 37°C, 5% CO2 in a humidified incubator for a post-exposure recovery time totaling ~18 hours for solid test article and controls. Tissue viability was assessed by 3-(4,5-dimethylthiazol -2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.
- MTT Auto reduction and colouring assessment
- MTT Pre-test
~50 mg of test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione was added to 1 mL of MTT media (~1 mg/mL) and incubated in a humidified incubator at approximately 37°C and approximately 5% CO2 for 2 hours, 55 minutes and 02 seconds. 50 µL of ultrapure water was used as a negative control.
- Test Article Color Test
Approximately 50 mg of test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione was added to 1.0 mL of ultrapure water and 2.0 mL isopropanol and incubated in a humidified incubator at approximately 37°C and approximately 5% CO2 for 2 hours, 04 minutes and 35 seconds. Samples were then added to the wells of a clear 96-well plate and the plate was read on a BioTek Synergy H4 (or equivalent) plate reader set to 570 nm. Test articles that tested positive for excessive coloration (OD >0.08) were assessed on living-tissue controls that were incubated in both culture media and MTT media as well (n=3 for both conditions).
- MTT Assay
After the recovery period, the MTT assay was performed on run 1 tissues by transferring the tissues to 24-well plates containing 300 µL MTT medium (1.0 mg/mL). After 2 hours 55 minutes and 00 seconds (solids) of MTT incubation at approximately 37°C, approximately 5% CO2 in a humidified incubator, the tissues were rinsed twice with DPBS. The blue formazan salt was extracted by submerging tissues in 2 mL isopropanol in a 24-well plate. The extraction time was 2 hours 00 minutes with gentle shaking for solid exposed tissues. The optical density of the extracted formazan (200 µL/well of a 96-well plate) was determined using a Synergy H4 spectrophotometer at 570 nm. Relative cell viability was calculated for each tissue as % of the mean negative control tissues.
- Evaluation of Test Article in the cell Models
1. Cell System:
Upon receipt, the MatTek EpiOcular™ tissue cultures were placed in 1.0 mL of fresh Maintenance medium (in a 6-well plate) for 59 minutes and 00 seconds. The tissues were not incubated overnight.
Control and Test Article Exposures:
20 µL of calcium and magnesium free DPBS was added to each tissue and the tissues placed back into the incubator for 31 minutes 00 seconds. The controls and the test article will be applied topically to tissues by pipette. Three tissues will be used per test compound and control.
a)Controls:
50 µL of negative control sterile ultrapure water, positive control methyl acetate were added to the tissues. The tissues were placed into the ~37°C humidified incubator with 5% CO2 for the approximately 6 hour exposure time.
b)Test Article:
Approximately 50 mg of the test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione were added to the tissues as a fine powder. The tissues were placed into the ~37°C humidified incubator with 5% CO2 for the approximately 6 hour exposure time.
3. Post exposure treatment
After the exposure, the tissues were rinsed 20 to 25 times with ~1 mL of DPBS to remove test material. The apical surface was gently blotted with a cotton swab and cultures were immediately transferred to a 12-well plate containing 5 mL of media per well. Tissues exposed to solid test article (and the respective control) were incubated, submerged in the media for ~25 minutes at room temperature. Tissuses were then transferred to 6-well plates containing 1.0 mL fresh Maintenance medium per well and incubated for a post-exposure recovery period for 17 hours 48 minutes 01 seconds at approximately 37 degC, 5% CO2 in a humidified incubator.
- Doses of test chemical and control substances used
a)Test Article:
Approximately 50 mg of 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione were added to the tissues as a fine powder. The tissue s were placed into the ~37°C humidified incubator with 5% CO2 for the approximately 6 hour exposure time.
b)Controls:
50 µL of negative control sterile ultrapure water, positive control methyl acetate were added to the tissues. The tissues were placed into the ~37°C humidified incubator with 5% CO2 for the approximately 6 hour exposure time.
- Duration and temperature of exposure, post-exposure immersion and post-exposure incubation periods:
Tissues were exposed for approximately ~6 hours for test article and controls, at approximately 37°C, 5% CO2 in a humidified incubator.
Following the post soak, the tissues were rinsed and incubated at approximately 37°C, 5% CO2 in a humidified incubator for a post-exposure recovery time totaling ~18 hours for test article and controls.
- Justification for the use of a different negative control than ultrapure H2O (Not applicable)
- Justification for the use of a different positive control than neat methyl acetate (Not applicable)
- Number of tissue replicates used per test chemical and controls: 3 tissues were used for test compound and control.
- Description of the method used to quantify MTT formazan
The blue formazan salt was extracted by submerging tissues in 2 mL isopropanol in a 24-well plate. The extraction time was 2 hours 00 minutes with gentle shaking for solid exposed tissues. The optical density of the extracted formazan (200 µL/well of a 96-well plate) was determined using a Synergy H4 spectrophotometer at 570 nm.
- Description of evaluation criteria used including the justification for the selection of the cut-off point for the prediction model
Calculations and Statistical Methods
MTT Assay
Blanks:
· The OD mean from all replicates for each plate (ODblank).
Negative Controls (NC):
· The blank corrected value was calculated: ODNC= ODNCraw– ODblank.
· The OD mean per NC tissue was calculated.
· The mean OD for all tissues corresponds to 100% viability.
· The mean, standard deviation (SD), standard error of the mean (SEM) and the percent coefficient of variation (% CV) was calculated.
ODblank= optical density of blank samples (isopropanol alone).
ODNCraw= optical density negative control samples.
ODNC= optical density of negative control samples after background subtraction.
Positive Control (PC):
· Calculate the blank corrected value: ODPC= ODPCraw– ODblank.
· The OD mean per PC tissue was calculated.
· The viability per tissue was calculated: %PC = [ODPC/ mean ODNC] x 100.
· The mean viability for all tissues was calculated: Mean PC = Σ %PC / number of tissues.
· The standard deviation (SD), standard error of the mean (SEM) and the percent coefficient of variation (% CV) was calculated.
ODPCraw= optical density positive control samples.
ODPC= optical density of positive control samples after background subtraction.
Tested Articles:
· Calculate the blank corrected value ODTT= ODTTraw– ODblank.
· The OD mean per tissue is calculated.
· The viability per tissue is calculated: %TT = [ODTT/ mean ODNC] x 100.
· The mean viability for all tissues is calculated: Mean TT = Σ %TT / number of tissues.
· The standard deviation (SD) and the percent coefficient of variation (% CV)for the controls and the test articles will be calculated.
ODTTraw= optical density test article samples.
ODPC= optical density of test article samples after background subtraction.
Data Correction Procedure for MTT Interfering Compounds
True viability = Viability of treated tissue – Interference from test article = ODtvt – ODkt where ODkt = (mean ODtkt – mean ODukt).
ODtvt = optical density of treated viable tissue
ODkt = optical density of killed tissues
ODtkt = optical density of treated killed tissue
ODukt = optical density of untreated killed tissue (NC treated tissue)
Data Correction Procedure for Colored Compounds
True viability = Viability of treated tissue incubated in MTT media – Viability of treated tissue incubated in media without MTT = ODtvt – ODvt.
ODtvt = optical density of treated viable tissue incubated in MTT media
ODvt = optical density of viable tissues incubated in media alone.
Proposed Statistical methods
The mean, standard deviation (SD) and the percent coefficient of variation (% CV) for the controls and the test article will be calculated.
- Evaluation of data
The results of the assay was evaluated and compared to negative control.
Table: Irritancy Prediction
In VitroResults In VivoPrediction
Mean tissue viability ≤60% Irritant (I) – Category 1 or 2
Mean tissue viability >60% Non-irritant (NI) – No Category
- Assay quality controls
- Negative Controls (NC)
The assay is meeting the acceptance criterion if the mean viability of the NC in terms of Optical Density (OD570) of the NC tissues (treated with sterile ultrapure water) in the MTT assay are >0.8 to <2.5. This is an indicator of tissue viability following shipping and conditions under use.
- Positive Controls (PC)
Methyl acetate was used as a PC and tested concurrently with the test article. The assay is meeting the acceptance criteria if the viability of the PC is <50% of the negative control.
- Standard Deviation (SD)
Each test of ocular irritancy potential is predicted from the mean viability determined on 3 single tissues. The assay meets the acceptance criteria if SD calculated from individual percent tissue viabilities of the replicates is <18% for three replicate tissues.
- Irritation parameter:
- other: mean % tissue viability
- Run / experiment:
- Run 1
- Value:
- 91.5
- Vehicle controls validity:
- not specified
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- no indication of irritation
- Other effects / acceptance of results:
- The MTT data show the assay quality controls were met, as the OD of the negative control tissues was between 0.981 and 1.284 in run 1. Also, the positive control, methyl acetate, reduced tissue viability to be 11.3% of negative control (for 6 hour exposures with solids) in run one and the standard deviation (SD) calculated from individual percent tissue viabilities of the test article exposed replicates was 5.0 passing the acceptance criteria.
- Interpretation of results:
- other: not irritating
- Conclusions:
- The ocular irritation potential of test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined according to the OECD 492 test guideline followed for this study. The mean % tissue viability of test substance 2-octadecyl -1H-thioxantheno [2,1,9- def]isoquinoline-1,3(2H)-dione was determined to be 91.5%. Thus, substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) is considered to be "not irritating" to the human eye.
- Executive summary:
The ocular irritation potential of test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined according to the OECD 492 test guideline followed for this study. The MatTek EpiOcular™ model was used to assess the potential ocular irritation of the test articles by determining the viability of the tissues following exposure to the test article via MTT. The objective of this study was to assess the ocular irritation potential of test article. Tissues were exposed to test article and controls for ~6 hours, followed by a ~25 minute post-soak and approximately 18 hour recovery after the post-soak. The viability of each tissue was determined by MTT assay.
The MTT data show the assay quality controls were met, as the OD of the negative control tissues was between 0.981 and 1.284 in run 1. Also, the positive control, methyl acetate, reduced tissue viability to be 11.3% of negative control (for 6 hour exposures with solids) in run one and the standard deviation (SD) calculated from individual percent tissue viabilities of the test article exposed replicates was 5.0 passing the acceptance criteria.
The mean % tissue viability of test substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined to be 91.5%.
Hence, under the experimental test condition it was concluded that test substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) is considered to be not irritating to the human eye and being classified as “Not classified’’ as per CLP Regulation.
Reference
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (not irritating)
Respiratory irritation
Endpoint conclusion
- Endpoint conclusion:
- no study available
Additional information
Skin irritation
In different studies, 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) has been investigated for potential for dermal irritation to a greater or lesser extent. The studies are based on in vitro and in vivo experiments in rabbits along with human data for target chemical and its structurally similar read across substances, Acequinocyl(CAS No: -57960-19-7)
The dermal irritation potential of test article 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined according to the OECD 439 test guideline followed for this study. The MatTek EpiDerm™ model was used to assess the potential dermal irritation of the test article by determining the viability of the tissues following exposure to the test article via MTT. The objective of this study was to assess the dermal irritation potential of test article. Tissues were exposed to test article and controls for ~one hour, followed by a 42 hour post-exposure recovery period. The viability of each tissue was determined by MTT assay.
The MTT data show the assay quality controls were met, as the OD of the negative control tissues was between 1.195 and 1.430. Also, the positive control, 5% sodium dodecyl sulfate (SDS), reduced tissue viability to 4.5% of negative control and the standard deviation (SD) calculated from individual percent tissue viabilities of the test article exposed replicates was 2.9 passing the acceptance criteria.
The Mean % tissue viability compared to negative control (n=3) of the test substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione was determined to be 103.5 %.
In other study by (The rapporteur Member State the Netherlands, 2005) with similar substance (57960-19-7) skin irritation was observed.The skin irritation studyof Acequinocyl(CAS No: -57960-19-7)was performed on 6 female rabbits under an occlusion for observation period of 72 hours.
The test sample wasapplied to the skin of rabbits at a dose of 0.5 ml for 4 hours exposure period and scores were observed after1, 24, 48 and 72 hours. No irritation was noted during the 72 hours observation period Therefore the test materiall Acequinocyl was found to be non irritant on rabbit’s skin.
In other study by (European chemical agency (ECHA), 2010) with similar substance (57960-19-7) was observed.The skin irritation study of Acequinocyl(CAS No: -57960-19-7)was performed on rabbits under an occlusion for observation period of 72 hours. The test sample was applied to the skin of rabbits and scores were observed after1, 24, 48 and 72 hours. No irritation was observed during the 72 hours observation period Therefore the test material Acequinocyl was found to be non irritant on rabbit’s skin.
On the basis of available information for the target as well as read across substance , the test substance can be considered as not irritating to the skin.
Eye Irritation:
In different studies, 2-octadecyl-1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) has been investigated for potential for occular irritation to a greater or lesser extent. The studies are based on in vitro and in vivo experiments in rabbits along with human data for target chemical and its structurally similar read across substances, Acequinocyl(CAS No: -57960-19-7)
The ocular irritation potential of test article 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined according to the OECD 492 test guideline followed for this study. The MatTek EpiOcular™ model was used to assess the potential ocular irritation of the test articles by determining the viability of the tissues following exposure to the test article via MTT. The objective of this study was to assess the ocular irritation potential of test article. Tissues were exposed to test article and controls for ~6 hours, followed by a ~25 minute post-soak and approximately 18 hour recovery after the post-soak. The viability of each tissue was determined by MTT assay.
The MTT data show the assay quality controls were met, as the OD of the negative control tissues was between 0.981 and 1.284 in run 1. Also, the positive control, methyl acetate, reduced tissue viability to be 11.3% of negative control (for 6 hour exposures with solids) in run one and the standard deviation (SD) calculated from individual percent tissue viabilities of the test article exposed replicates was 5.0 passing the acceptance criteria.
The mean % tissue viability of test substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) was determined to be 91.5%.
Hence, under the experimental test condition it was concluded that test substance 2-octadecyl -1H-thioxantheno[2,1,9-def]isoquinoline-1,3(2H)-dione (CAS No.- 27870-92-4) is considered to be not irritating to the human eye and being classified as “Not classified’’ as per CLP Regulation.
Studies on similar substance 57960-19-7, from Draft Assessment Report (DAR), July 2005, no irritation was observed in rabbits. An eye irritation study in rabbits was performed in accordance with OECD 405. The treated eyes were washed 24 hours after instillation of the test substance in the ‘unwashed group’ (6 rabbits) and in the ‘washed group’ (3 rabbits) the treated eyes were washed 2 to 3 minutes after application. According to OECD 405 an eye wash may be executed 24 hour after application of a solid test substance. Therefore, only the ‘unwashed’ group was treated according to OECD 405, hence only the results of this group are reported. After 24 h only slight conjunctival redness was observed in 3 out of 6 rabbits. Acequinocyl was considered to be not irritating to rabbit eyes.
Therefore, on the basis of available information for the target as well as its read across, the test chemical 2-Octadecyl-1H-thioxantheno(2,1,9-def)isoquinoline-1,3(2H)dione was considered to be a non irritant to eyes.
Justification for selection of skin
irritation / corrosion endpoint:
The test substance Solvent Yellow
98 is determined to be not irritating when tested in accordance with
OECD guideline 439
Justification for selection of eye irritation endpoint:
The test substance Solvent Yellow 98 is determined to be not irritating when tested in accordance with OECD guideline 493
Justification for classification or non-classification
The test substance Solvent yellow 98 was classified as non –irritant to both skin and eye.
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