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EC number: 240-357-1 | CAS number: 16245-77-5
- 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:
- From 5 March 2018
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- Data is from experimental study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 018
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Version / remarks:
- 28 July 2015
- Deviations:
- yes
- Remarks:
- The living tissues treated with the test item were applied less than 8 mg of test item. As it was noted that all tissues were evenly covered by the test item, this deviation was not considered to comprise the outcome of the study.
- Principles of method if other than guideline:
- To evaluate the skin irritation potential of the test item Benzene-1,4-diammonium sulphate (CAS No:16245-77-5) /E507469 using the EpiskinTM reconstructed human epidermis model.
- GLP compliance:
- yes (incl. QA statement)
Test material
- Reference substance name:
- Benzene-1,4-diammonium sulphate
- EC Number:
- 240-357-1
- EC Name:
- Benzene-1,4-diammonium sulphate
- Cas Number:
- 16245-77-5
- Molecular formula:
- C6H8N2.H2O4S
- IUPAC Name:
- benzene-1,4-diammonium sulphate
- Test material form:
- solid: particulate/powder
Constituent 1
- Specific details on test material used for the study:
- SOURCE OF TEST MATERIAL- Source and lot/batch No.of test material: supplied by the sponsor, batch no. 60810- Expiration date of the lot/batch: 09 November 2018- Purity test date:20 July 2017STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL- Storage condition of test material: At room temperature, protected from humidity- Stability under test conditions:not applicable- Solubility and stability of the test substance in the solvent/vehicle: not applicable- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not applicableTREATMENT OF TEST MATERIAL PRIOR TO TESTING- Treatment of test material prior to testing: the test item was not treated before testing and was used neat- Preliminary purification step (if any): not applicable- Final dilution of a dissolved solid, stock liquid or gel: not applicable- Final preparation of a solid: no final preparationFORM AS APPLIED IN THE TEST (if different from that of starting material) neat
In vitro test system
- Test system:
- human skin model
- Source species:
- human
- Cell type:
- other: Human reconstructed epidermis
- Cell source:
- other: not applicable
- Source strain:
- other: not applicable
- Justification for test system used:
- Based on the peer review of the results of an inter-laboratory study with the EpiskinTM model, the ECVAM Scientific Advisory Committee (ESAC) endorsed the conclusion that the EpiskinTM model can be used for distinguishing between skin irritant and non-irritant chemicals.
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE- Model used: EpiskinTM- Tissue batch number(s):Not specified- Production date: not specified- Shipping date: not specified- Delivery date:not specified- Date of initiation of testing: 5 March 2018TEMPERATURE USED FOR TEST SYSTEM- Temperature used during treatment / exposure: 37°C- Temperature of post-treatment incubation (if applicable): 27°CREMOVAL OF TEST MATERIAL AND CONTROLS-Volume and number of washing steps: At the end of the treatment period, each tissue was removed from the well of the treatment plate, and rinsed with D-PBS. Rinsing was achieved by gently filling and emptying several times each tissue with D PBS to gently remove any residual test or control items.- Observable damage in the tissue due to washing: No damage- Modifications to validated SOP: No modificationsMTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE- MTT concentration: 0.3 mg/mL- Incubation time: 3 hours (±5 minutes) - Spectrophotometer: plate reader- Wavelength: 570 nm- Filter: no filter was used- Filter bandwidth: no filter was used- Linear OD range of spectrophotometer: not specifiedFUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATAhistorical data were not mentioned in the reportNUMBER OF REPLICATE TISSUES: triplicates per condition were used CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE- water-killed tissues- Procedure used to prepare the killed tissues (if applicable): untreated EpiskinTM tissues were placed in a 12 well plate containing 2 mL of water for injections per well. The 12-well plate was placed in an incubator (set at +37°C, 5% CO2) for 48 hours (± 1 hour). At the end of the incubation period, the tissues were dried before being placed in a freezer (-20°C) for at least one night. Once killed, the tissues were stored in the freezer for up to 6 months. Before using, the water-killed tissues were thawed at room temperature (for at least 1 hour ± 5 minutes in 2 mL of maintenance medium).- N. of replicates : triplicates were used- Method of calculation used:-Non-Specific MTT reduction (NSMTT) calculation:-cOD Untreated Killed tissues = ODUK - mean ODblank,-cOD Test Item-treated Killed tissues = ODTIK - mean ODblank.NSMTT = [(mean cODTIK - mean cODUK) / mean cODNC] x 100True MTT metabolic conversion (TODTI):- TODTI = [mean cODTI - (mean cODTIK - mean cODUK)].Then, the relative mean viability is calculated as follows:Relative mean viability = (TODTI / mean cODNC) x 100NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: Two independent sequences : pretest and main test QUANTIFICATION OF IL-1 AlphaThe concentration of the IL-1α in the culture medium retained following the 42-hour recovery period was determined only for test items found with a mean relative viability > 50% after the MTT reduction. For these test items, culture media samples retained from the three negative controls and the three test item-treated tissues were analyzed by ELISA according to our internal procedures. The assay for the determination of IL-1α concentration in EpiskinTM culture medium is an ELISA (Enzyme Linked Immunosorbent Assay) in which anti-IL-1α antibodies are coated in wells of a 96 wells plate. The IL 1α present in the tested samples bind to the immobilised anti-IL-1α antibodies and are revealed using a peroxidase (HRP) conjugated anti-IL-1α antibody.Enzymatic reaction is performed by using a substrate which is transformed into a colored product, which intensity is measured in OD value using a spectrophotometer. The analyte concentration concentrations can be then determined by interpolation from a calibration curve in which OD values are plotted against IL 1α concentrations of calibration standards. PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)- The test substance is considered to be irritant to skin if the viability after 15 minutes exposure and 42 hours post exposure period is less than or equal to 50% or higher than 50M associated with a Il-1 alpha higher than 60 pg/mL- The test substance is considered to be non-corrosive to skin if the viability after 15 minutes exposure and 42 hours post exposure period is greater 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):10 mg ± 2mg- Concentration (if solution): pureVEHICLENo vehicle was usedNEGATIVE CONTROL- Amount(s) applied (volume or weight): 10 µL- Concentration (if solution): neatPOSITIVE CONTROL- Amount(s) applied (volume or weight): 10µL- Concentration (if solution): 5% (w/v)
- Duration of treatment / exposure:
- 15 minutes
- Duration of post-treatment incubation (if applicable):
- 42 hours
- Number of replicates:
- triplicates
Results and discussion
In vitro
Resultsopen allclose all
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- Test Item (true MTT metabolic conversion)
- Value:
- 94
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- other: the IL-1α concentrations (pg/mL)
- Run / experiment:
- Test item
- Value:
- < 5
- Negative controls validity:
- valid
- Other effects / acceptance of results:
- - OTHER EFFECTS:- Visible damage on test system: no damage- Direct-MTT reduction: The MTT solution containing the test item turned blue/purple when compared with the negative control. The test item was therefore considered to have direct MTT reducing properties. As a result, additional controls were performed on water-killed tissues in parallel to the main test.- Colour interference with MTT: During this test, as the water solution containing the test item did not change colour, the test item was found not to have a colouring potential. As a result, no additional controls were used in parallel to the main test.DEMONSTRATION OF TECHNICAL PROFICIENCY: Positive control condition showed proficiency of the test system to quantify tissue viabilityACCEPTANCE OF RESULTS: - Acceptance criteria met for negative control: yes- Acceptance criteria met for positive control: yes- Acceptance criteria met for variability between replicate measurements: yes-
Any other information on results incl. tables
Table 1 :Results
cOD |
| Viability(%) | NSMTT | ||
Groupe | Mean | SD | Mean | SD |
|
Negativecontrol | 0.738 | 0.031 | 100 | 4 |
|
positive control | 0.026 | 0.004 | 4 | 1 |
|
test item | 0.739 | 0.085 | 100 | 12 |
|
test item (deadtissues) | 0.079 | 0.013 | 11 | 2 | 6 |
untreated(deadtissues) | 0.036 | 0.015 | 5 | 2 |
|
Test item (true MTT metabolic conversion) | 0.697 |
| 94 |
|
|
cOD=blankcorrectedopticaldensity
MTT = « -(4,5-dimethylthiazol-2-ylà-2,5-diphenyltetrazoliumbromide
NSMTT= nonspecificMTTreduction
SD = standard deviation
Applicant's summary and conclusion
- Interpretation of results:
- GHS criteria not met
- Conclusions:
- Under the experimental condition of the study, the test item did not induced decrease of tissue viability (94%) and no IL1-alpha release in medium (under 5pg/mL) when applied on EpiskinTM. Hence, , the test item, E507469, is considered to be non-irritant to skin.According to the results of this study, the classification of the test item should be: No Category (UN GHS and Regulation (EC) No. 1272/2008).
- Executive summary:
The objective of this study was to evaluate the skin irritation potential of the test item, E507469, using the EpiskinTM reconstructed human epidermis model.
The study design was based upon the international guideline OECD No. 439. The study was conducted in compliance with the principles of Good Laboratory Practice.
Preliminary tests were performed to detect the ability of the test item to directly reduce MTT as well as its colouring potential. Following the preliminary tests, the skin irritation potential of the test item was tested in the main test. The test item and both the negative and positive controls were applied topically on triplicate tissues and incubated at room temperature for 15 minutes. At the end of the treatment period, each tissue was rinsed with D-PBS and incubated for 42 hours at +37°C, 5% CO2 in a humidified incubator. The cell viability was then assessed by means of the colourimetric MTT reduction assay. Relative viability values were calculated for each tissue and expressed as a percentage of the mean viability of the negative control tissues which was set at 100% (as reference viability).
In addition, the concentration of the inflammatory mediator IL-1α was evaluated in the culture medium retained following the 42-hour recovery period. This quantification, based on an ELISA assay, was performed since the mean relative viability of the test item-treated tissues was > 50% following the MTT reduction assay.
In the preliminary tests, the test item was found to have direct MTT reducing properties but no colouring properties. All acceptance criteria for the negative and positive controls were fulfilled. The study was therefore considered to be valid.
Following a 15 minutes exposure and 42 hours of recovery period, the relative mean viability of the tissues treated with the test item was 94%.
As the mean relative viability > 50% after the MTT reduction, the IL-1α concentrations in culture media samples retained from the three negative controls and the three test item-treated tissues were analyzed by ELISA. The IL-1α concentration value of two tissues was found below the limit of quantification (< 5.00 pg/mL). Consequently, the mean IL-1α concentration from the three test item-treated tissues was not calculated. The IL-1α concentration value of the other test item-treated tissue was found < 60 pg/mL (11.2 pg/mL). Therefore, the results met the criteria for an in vitro classification as non-irritant to skin.
Under the experimental conditions of this study, the test item, E507469, is considered to be non-irritant to skin.
According to the results of this study, the classification of the test item should be: No Category (UN GHS and Regulation (EC) No. 1272/2008).
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