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EC number: 201-744-0 | CAS number: 87-41-2
- 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 vivo
Administrative data
- Endpoint:
- in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Guideline study (OECD 474); GLP compliant
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 005
- Report date:
- 2005
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 870.5395 (In Vivo Mammalian Cytogenetics Tests: Erythrocyte Micronucleus Assay)
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- micronucleus assay
Test material
- Reference substance name:
- Phthalide
- EC Number:
- 201-744-0
- EC Name:
- Phthalide
- Cas Number:
- 87-41-2
- Molecular formula:
- C8H6O2
- IUPAC Name:
- 1(3H)-Isobenzofuranone
- Details on test material:
- - Name of test material (as cited in study report): Phthalide
- Physical state: Solid, white
- Analytical purity: 100%
- Lot/batch No.: S12233-025
- Stability under test conditions: The stability of the test substance under storage conditions throughout the study period is guaranteed.
- Storage condition of test material: room temperature
Constituent 1
Test animals
- Species:
- mouse
- Strain:
- NMRI
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River GmbH, Germany.
- Age at study initiation: 5 - 8 weeks.
- Weight at study initiation: 27 g (mean).
- Assigned to test groups randomly: yes, under following basis: plan prepared with an appropriate computer program.
- Housing: individually in Makrolon cages.
- Diet: Standardized pelleted feed (Maus/Rafte Haltung "GLP", Provimi Kliba SA, Kaiseraugst, Switzerland), ad libitum.
- Water: tap water, ad libitum.
- Acclimation period: 5 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24 °C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12/12
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- DMSO is used up to 4 ml/kg body weight only, the DMSO solution was filled up to 10 ml/kg body weight with olive oil and mixed thoroughly.
- Duration of treatment / exposure:
- The animals were treated once and samples of bone marrow were taken 24 hours and 48 hours after the treatment.
- Frequency of treatment:
- single application.
- Post exposure period:
- 24 hours and 48 hours
Doses / concentrations
- Remarks:
- Doses / Concentrations:
187.5, 375.0 and 750.0 mg/kg bw
Basis:
nominal conc.
- No. of animals per sex per dose:
- 5
- Control animals:
- yes, concurrent vehicle
- Positive control(s):
- Cyclophosphamide (CPP) / Vincristine Sulphate (VCR)
- Route of administration: CPP: orally/ VCR: intraperitoneally
- Doses / concentrations: CPP: 20 mg / VCR: 0.15 mg
Examinations
- Tissues and cell types examined:
- bone marrow
- Details of tissue and slide preparation:
- CRITERIA FOR DOSE SELECTION: Pretests were performed
DETAILS OF SLIDE PREPARATION:
The bone marrow was prepared according to the method described by SCHMID, W. and SALAMONE, M. et al.
• The two femora of the animals sacrificed by cervical dislocation were prepared by dissection and removing all soft tissues.
• After cutting off the epiphyses, the bone marrow was flushed out of the diaphysis into a centrifuge tube using a cannula filled with fetal calf serum (FCS) which was at 37°C (about 2 mL/femur).
• The suspension was mixed thoroughly with a pipette, centrifuged at 300 x g for 5 minutes, the supernatant was removed and the precipitate was resuspended in about 50 µL fresh FCS.
• 1 drop of this suspension was dropped onto clean microscopic slides, using a Pasteur pipette. Smears were prepared using slides with ground edges, the preparations were dried in the air and subsequently stained.
• The slides were stained in eosin and methylene blue (modified May-Gruenwald solution or Wrights solution) for about 5 minutes.
• After having briefly been rinsed in purified water, the preparations were soaked in purified water for about 2 - 3 minutes.
• Subsequently, the slides were stained in Giemsa solution (15 mL Giemsa, 185 mL purified water) for about 15 minutes.
• After having been rinsed twice in purified water and clarified in xylene, the preparations were mounted in Corbit-Balsam. - Evaluation criteria:
- A finding is considered positive if the following criteria are met:
• Significant and dose-related increase in the number of PCEs containing micronuclei.
• The number of PCEs containing micronuclei has to exceed both the concurrent negative control and the highest value of the historical control range.
A test substance is considered negative if the following criteria are met:
• The number of cells containing micronuclei in the dose groups is not significantly above the negative control and is within the historical control data. - Statistics:
- The statistical evaluation of the data was carried out using the program system MUKERN (BASF Aktiengesellschaft).
The asymptotic U test according to MANN-WHITNEY (modified rank test according to WILCOXON) was carried out to clarify the question whether there were significant differences between the control group and dose groups with regard to the micronucleus rate in polychromatic erythrocytes. The relative frequencies of cells containing micronuclei of each animal were used as a criterion for the rank determination for the U test. Significances were identified as follows:
* p ≤ 0.05
** p ≤ 0.01
Results and discussion
Test results
- Sex:
- male/female
- Genotoxicity:
- negative
- Toxicity:
- yes
- Remarks:
- poor general state
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
Any other information on results incl. tables
Summary of Micronucleus Test Results:
Dose: mg/kg bw | Total No. PCE´s | NCE´s/PCE´s | Micronuclei in PCE´s (‰) |
Micronuclei in NCE´s (‰) |
|
DMSO (24 h) | 20000 | 7161 | 1.6 | 0.8 | |
DMSO (48 h) | 20000 | 10024 | 1.9 | 1.6 | |
187.5 (24 h) | 20000 | 7084 | 2.5* | 2.3 | |
375 (48 h) | 20000 | 6441 | 2.1 | 1.6 | |
750 (24 h) | 20000 | 6930 | 1.9 | 0.7 | |
750 (48 h) | 20000 | 11208 | 2.2 | 1.4 | |
20 mg/kg bw Cyclophosphamide (24 h) | 10000 | 3923 | 16.9** | 0.8 | |
0.15 mg/kg bw Vincristine (24 h) | 10000 | 5030 | 58.7** | 1.4 |
WILCOXON TEST (ONE-SIDED) : *: p <= 0 .05, **: p <= 0.0 1
According to the results of the present study, there are thus no biologically relevant, significant differences in the frequency of erythrocytes containing micronuclei either between the vehicle control and the 3 dose groups (187.5 mg/kg, 375.0 mg/kg and 750.0 mg/kg) or between the two sacrifice intervals (24 and 48 hours).
The number of normochromatic or polychromatic erythrocytes containing small micronuclei (d < D/4) or large micronuclei (d > D/4) did not deviate from the vehicle control value at any of the sacrifice intervals and was within the historical control range. No inhibition of erythropoiesis determined from the ratio of polychromatic to normochromatic erythrocytes was detected .
The statistically significance (p< 0.05) observed at the lowest dose of 187.5 mg/kg body weight after a sacrifice interval of 24 hours is without any biological relevance; a value of 2.5‰ micronucleated polychromatic erythrocytes is well within the historical control range and there is not any dose-response relationship.
Under the experimental conditions chosen here, the test substance phthalide did not lead to any increase in the number of polychromatic erythrocytes containing either small or large micronuclei, thus has no chromosome-damaging (clastogenic) effect nor does it lead to any impairment of chromosome distribution in the course of mitosis (aneugenic activity) in bone marrow cells in vivo.
Applicant's summary and conclusion
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