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EC number: 425-180-1 | CAS number: 66170-10-3
- 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
Genetic toxicity: in vitro
Administrative data
- Endpoint:
- in vitro gene mutation study in mammalian cells
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- migrated information: read-across from supporting substance (structural analogue or surrogate)
- Adequacy of study:
- key study
- Study period:
- 1981
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Not guideline conformant, GLP: no data
Data source
Reference
- Reference Type:
- publication
- Title:
- Unnamed
- Year:
- 1 981
- Report date:
- 1981
Materials and methods
Test guideline
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)
- Deviations:
- not applicable
- Principles of method if other than guideline:
- Method: other: according to Amacher, D.E. et al.: Mutat. Res. 64, 391-406
- GLP compliance:
- not specified
- Type of assay:
- mammalian cell gene mutation assay
Test material
- Reference substance name:
- Ascorbic acid
- EC Number:
- 200-066-2
- EC Name:
- Ascorbic acid
- Cas Number:
- 50-81-7
- IUPAC Name:
- 5-(1,2-dihydroxyethyl)-3,4-dihydroxyfuran-2(5H)-one (non-preferred name)
- Details on test material:
- - Name of test material : Ascorbic acid (in parallel sodium ascorbate was applied)
- Analytical purity: 99.9%
Constituent 1
Method
- Target gene:
- Thymidine Kinase (TK)
Species / strain
- Details on mammalian cell type (if applicable):
- L5178Y mouse lymphoma cells used in these experiments were the 3.7.2C line and were negative when screened for mycoplasma via growth on agar and Hoechst staining methods. Weekly treatment of these cells to reduce the TK-/- background mutant frequencies were performed. Culture medium was RPM1-1640 (Microbiological Associates) containing 50 units/ml penicillin-50 µg/mL streptomycin, 0.05 % Pluranic F-68 (BASF Wyandotte), and either 3 % horse serum (R3 or test medium) or 5 % horse serum (R5 or growth medium).
- Additional strain / cell type characteristics:
- not specified
- Metabolic activation:
- without
- Test concentrations with justification for top dose:
- up to ca. 704 µg/mL (4 mM; ascorbic acid); up to ca. 475 µg/mL (2.4 mM; sodium ascorbate)
- Vehicle / solvent:
- saline (1%)
Controls
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- ethylmethanesulphonate
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in medium
DURATION
- Preincubation period: 1 h
- Exposure duration: 3 h
- Expression time (cells in growth medium): 48 h
The dosing of cells, removal of test chemicals after 3-h exposure, and maintenance of cells in lag phase growth for the 48-h expression period were previously described [Armacher, D.E. et al., 1979, Mutat. res., 64, 391-406]. Individual test cultures contained 6 x E06 cells in R3 medium, ascorbate or ascorbic acid initially dissolved in saline, and in same cases, 0.1 mg/mL freshly prepared catalase. Final solvent (saline) concentration was always 1%. Negative controls received saline only; concurrent
positive controls were dosed with 2.5 X 10-3 M ethylmethanesulfonate (EMS) dissolved in saline and filter-sterilized. In one set of experiments, ascorbate in R3 medium with or without catalase was preincubated for 1 h at 37°C prior to the addition of cells for 3-h exposure. Same of these preincubated samples were heated to 56°C for 15 min then coaled before cells were added. Where indicated, pH was measured before the 3-h incubation period with a Copenhagen Radiometer an duplicate cultures prepared for that purpose (sterility cannot be maintained after pH measurements). Test chemicals were removed by repeated washing and cells maintained at 37°C at cell densities permitting log phase growth in a New Brunswick Roller-drum. Cloning in soft-agar medium containing 0.37 % Difco Noble agar, 10 % serum and no antibiotics. Viable colony counts and counts of large trifluarathymidine-resistant (TFTr) colonies were made after 7 days incubation af plates at 37 °C in a humid 5 % CO2 atmosphere via a calibrated Artek model 870. Cell survival was determined as the product of both daily growth and cloning efficiency values relative to saline controls. This method differs from the colony forming method used by Rosin et al. and is sensitive to minor day-to day fluctuations. - Evaluation criteria:
- Viable colony counts and counts of large trifluarathymidine-resistant (TFTr) colonies were made after 7 days incubation of plates at 37 °C in a humid 5 % CO2 atmosphere via a calibrated Artek model 870.
- Statistics:
- mean and standard deviation
Results and discussion
Test results
- Species / strain:
- mouse lymphoma L5178Y cells
- Metabolic activation:
- without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- ca. 264 µg/mL (1.5 mM; ascorbic acid); ca. 99 µg/mL (0.5 mM; sodium ascorbate)
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- Ascorbic acid as well as sodium ascorbate was tested. Ascorbic acid was tested at concentrations up to 4.0 mM (ca. 704 µg/mL). It was cytotoxic at concentrations above 1.5 mM (ca. 265 µg/mL) but not mutagenic up to the highest concentration used. Sodium ascorbate was tested at concentrations up to 2.4 mM (ca. 475 µg/ml). It was cytotoxic at concentrations above 0.5 mM (ca. 99 µg/ml) but not mutagenic up to the highest concentration used.
Mutagenicity for the concurrent positive control (2.5 X 10-3 M EMS) appears to vary considerably; however, the results represent pooled data from independent trials spanning a 6 months.
Very high levels of ascorbic acid exceed the buffering capacity of the tissue culture medium and cause a precipitous drop in pH. To determine what effect increased hydrogen ion concentration might have an observed mutagenicity and cytotoxicity, R3 medium was titrated with 1 N HCl, filtered, measured for pH, then added to cultures of L5178Y cells. A pH of less than 6 results in sharply increased cytotoxicity, but no dose-dependent increase in mutant frequency. Since none of the ascorbic acid concentrations produced an initial pH lower than 7, reduced pH was not responsible for ascorbic acid toxicity. Representative pH measurements of sodium ascorbate dissolved in R3 medium showed no initial change (pH 7.6-7.8) compared to untreated medium. - Remarks on result:
- other: strain/cell type: L5178Y
- Remarks:
- Migrated from field 'Test system'.
Applicant's summary and conclusion
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