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Diss Factsheets
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EC number: 212-379-1 | CAS number: 812-00-0
- 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:
- key study
- Study period:
- 2 October 2018 - 31 January 2019
- Reliability:
- 1 (reliable without restriction)
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 019
- Report date:
- 2019
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 442C (In Chemico Skin Sensitisation: Direct Peptide Reactivity Assay (DPRA))
- Deviations:
- yes
- Remarks:
- See Deviations text below.
- GLP compliance:
- yes (incl. QA statement)
- Type of study:
- other: DPRA
Test material
- Reference substance name:
- Methyl dihydrogen phosphate
- EC Number:
- 212-379-1
- EC Name:
- Methyl dihydrogen phosphate
- Cas Number:
- 812-00-0
- Molecular formula:
- CH5O4P
- IUPAC Name:
- methyl dihydrogen phosphate
- Test material form:
- liquid
Constituent 1
- Specific details on test material used for the study:
- Batch 34448
Purity 67.9 - 71.2% Methyl dihydrogen phosphate
Expiry 30 March 2022
In vitro test system
- Details on the study design:
- Preparation of test item
The concentration of the test item to be used was determined based on the molecular weight (MW) 96 g/mol and the purity, the purity value 67.9-71.2 % was treated as 67.9 %. The target weight (± 10 %) of the test item was calculated. 100 mM test item solution was prepared by dissolving 42.3 mg and 42.5 mg test item in 3 mL of the solvent acetonitrile for the Cys-peptide and Lys-peptide, respectively.
HPLC was used for the measurement of peptide depletion of the Cys-peptide and the Lys-Peptide.
Positive control
Positive controls were treated identically as the test item. The following positive controls were used:
• Cinnamaldehyde (CAS 104-55-2, food grade ≥95 %, batch no. MKBT8955V) was used as 100 mM (± 10 %) solution in acetonitrile for the Cys-peptide.
• 2,3-Butanedione (CAS 431-03-8, ≥99 %, batch no. BCBS3560V) was used as 100 mM (± 10 %) solution in acetonitrile for the Lys-peptide
As cinnamaldehyde mixed with the lysine peptide turned turbid in all experiments performed during the implementation phase, it was considered unsuitable as positive control. Instead, the proficiency chemical 2,3-Butanedione was used as positive control showing mid-range depletion for the lysine peptide.
Solvent controls
For both peptides, four sets of solvent controls using acetonitrile instead of test item stock solution were prepared in triplicate. Set A was analysed together with the peptide calibration standards, sets B1 and B2 were analysed at the start and end of the analysis sequence and were used as stability control for the peptide over the total analysis time. Set C was incubated and analysed together with the samples and was used for calculation of the peptide depletion.
Co-elution control
Sample prepared from the respective peptide buffer and the test item, but without peptide.
Test solutions
Dilution buffers
• 2 mL Acetonitrile were mixed with 8 mL phosphate buffer, pH 7.498 (Peptide dilution buffer C)
• 2 mL Acetonitrile were mixed with 8 mL ammonium acetate buffer, pH 10.204 (Peptide dilution buffer K)
Peptide stock solutions
The peptide stock solutions were freshly prepared for each assay.
• 0.667 mM Cys-Peptide solution was prepared by dissolving 20.0 mg of the peptide in 40 mL phosphate buffer, pH 7.498. (batch no. 20181024)
• 0.667 mM Lys-Peptide solution was prepared by dissolving 25.9 mg of the peptide in 50 mL ammonium acetate buffer, pH 10.204. (batch no.20181009)
Peptide calibration standards
From each peptide stock solution, the following calibration standards were prepared in the appropriate dilution buffer: 0.534 / 0.267 / 0.1335 / 0.0667 / 0.0334 / 0.0167 mM Peptide. Calibration samples were analysed before the samples containing the test item. Blank dilution buffer was also measured.
Test item samples
Samples were prepared in triplicate for each peptide. The Cys-peptide samples were prepared in 1:10 molar ratio (0.5 mM peptide: 5 mM test item), the Lys-peptide samples in 1:50 molar ratio (0.5 mM peptide and 25 mM test item) using the stock solutions. A final volume of 1 mL per sample was prepared for each sample.
Incubation
The positive control, solvent control sets C, and test item samples were incubated in closed amber glass HPLC vials in an incubation chamber at 25.0 °C for 22 h and 5 minutes for the Cys-peptide and 22 h for the Lys-peptide, respectively. All three replicates for the Cys-peptide were turbid after incubation. They were centrifuged (10 min, 400 g) and only the clear supernatant was used for the measurement. None of the replicates for the Lys-peptide were turbid after incubation.
According to the test guideline, the reactivity is classified as “high”, “moderate”, “low” or “minimal” using the Cysteine 1:10/Lysine 1:50 prediction model shown in the table below.
Results and discussion
- Positive control results:
- The mean peptide-depletion of the positive control 2,3 butanedione for the Lys-peptide was marginal out of the range of historical data, but nevertheless the value 40.25 % was within the range given in the OECD442C.
In vitro / in chemico
Resultsopen allclose all
- Key result
- Group:
- test chemical
- Run / experiment:
- mean
- Parameter:
- lysine depletion
- Value:
- 0 %
- Key result
- Group:
- test chemical
- Run / experiment:
- mean
- Parameter:
- cysteine depletion
- Value:
- 0 %
- Other effects / acceptance of results:
- No observations arousing doubts concerning the accuracy of the results and the validity of the study were made.
Any other information on results incl. tables
Calculated peptide depletion values for the Cys-Peptide
Sample name | Depletion (%) | ||
Single | Mean | SD | |
Positive control Rep 1 | 75.44 | 76.14 | 0.94 |
Positive control Rep 2 | 75.77 | ||
Positive control Rep 3 | 77.21 | ||
Test item Rep 1 | 0.00 (-1.07) | 0.00 | 0.00 |
Test item Rep 2 | 0.00 (-1.34) | ||
Test item Rep 3 | 0.00 (-1.23) |
Negative depletion values were considered as “zero” when calculating the mean.
Calculated peptide depletion values for the Lys-Peptide
Sample name | Depletion (%) | ||
Single | Mean | SD | |
Positive control Rep 1 | 38.14 | 40.25 | 3.99 |
Positive control Rep 2 | 37.76 | ||
Positive control Rep 3 | 48.46 | ||
Test item Rep 1 | 0.00 (-3.33) | 0.00 | 0.00 |
Test item Rep 2 | 0.00 (-0.73) | ||
Test item Rep 3 | 0.00 (-0.92) |
Negative depletion values were considered as “zero” when calculating the mean.
Applicant's summary and conclusion
- Conclusions:
- The mean peptide depletion in the Cys-peptide and Lys-peptide assay was 0.00 %, therefore the test item was classified with:
DPRA Prediction: Negative Reactivity class: Minimal - Executive summary:
DPRA Prediction: Negative Reactivity class: Minimal
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