Registration Dossier
Registration Dossier
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EC number: 701-008-3 | CAS number: -
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:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Guideline study (OECD), but not on test substance defined in Section 1.
- Justification for type of information:
- No in vivo studies are available on S278. Data from structurally related phthalates including DINP and BBP provide useful information on the genotoxicity potential of S278.
Cross-referenceopen allclose all
- Reason / purpose for cross-reference:
- read-across: supporting information
Reference
- Endpoint:
- in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- read-across from supporting substance (structural analogue or surrogate)
- Adequacy of study:
- key study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Guideline study (OECD), but not on test substance defined in Section 1.
- Justification for type of information:
- No in vivo studies are available on S278. Data from structurally related phthalates including DINP and BBP provide useful information on the genotoxicity potential of S278.
- Reason / purpose for cross-reference:
- read-across source
- Reason / purpose for cross-reference:
- reference to same study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
- Deviations:
- not specified
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)
- Deviations:
- not specified
- GLP compliance:
- not specified
- Type of assay:
- micronucleus assay
- Species:
- mouse
- Strain:
- CD-1
- Sex:
- male
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Laboratories, Portage, MI
- Age at study initiation: ca. 6-9 weeks
- Weight at study initiation: 17-35 g (although 17 g might include females used in another experiment)
- Assigned to test groups randomly: [no/yes, under following basis: ] no data
- Fasting period before study: no data
- Housing: no data
- Diet (e.g. ad libitum): no data
- Water (e.g. ad libitum): no data
- Acclimation period: no data
ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): no data
- Route of administration:
- oral: gavage
- Vehicle:
- - Vehicle(s)/solvent(s) used: vehicle control used, but identity not described in publication
- Details on exposure:
- no data
- Duration of treatment / exposure:
- Two consecutive days
- Frequency of treatment:
- Daily
- Post exposure period:
- One day
- Remarks:
- Doses / Concentrations:
500
Basis:
actual ingested
mg/kg bw/day - Remarks:
- Doses / Concentrations:
1000
Basis:
actual ingested
mg/kg bw/day - Remarks:
- Doses / Concentrations:
2000
Basis:
actual ingested
mg/kg bw/day - No. of animals per sex per dose:
- 5 males
- Control animals:
- yes, concurrent vehicle
- Positive control(s):
- cyclophosphamide
- Justification for choice of positive control(s): no data
- Route of administration: presumably gavage
- Doses / concentrations: 20 mg/kg bw/day - Tissues and cell types examined:
- Erythrocytes from bone marrow
- Details of tissue and slide preparation:
- CRITERIA FOR DOSE SELECTION:
No toxicity seen in a range-finding study at 2000 mg/kg bw/day.
TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
Treated on two consecutive days, and mice sacrificed on the third day.
DETAILS OF SLIDE PREPARATION:
Briefly, both femurs were removed from each treated mouse and the proximal ends were cut to expose the bone marrow, which was then aspirated with fetal bovine serum into a centrifuge tube. The cells were collected by centrifugation and slides were prepared. After fixation in methanol, the slides were stained with acridine orange for about 1-2 minutes.
METHOD OF ANALYSIS:
The slides were evaluated at 400x by fluorescence microscopy. A total of 1000 erythrocytes were counted from each animal, and the total numbers of polychromatic (PCE) and normochromatic (NCE) erythrocytes were tabulated. A total of 1000 PCEs were evaluated for the presence of micronuclei.
OTHER: - Evaluation criteria:
- no data
- Statistics:
- Statisitical analysis included means and standard deviations of the micronucleus test data, and a test of equality of means was performed by standard one-way analysis of variance.
- Sex:
- male
- Genotoxicity:
- negative
- Remarks:
- at 2000 mg/kg bw/day
- Toxicity:
- no effects
- Remarks:
- at 2000 mg/kg bw/day
- Vehicle controls validity:
- not specified
- Negative controls validity:
- not examined
- Positive controls validity:
- valid
- Additional information on results:
- RESULTS OF RANGE-FINDING STUDY
- Dose range: up to 2000 mg/kg bw/day (no details given of other dose levels)
- Solubility: no data
- Clinical signs of toxicity in test animals: no evidence of toxicity (extent of examination unclear from report)
- Evidence of cytotoxicity in tissue analyzed: no data
RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): Percentage PCEs with micronuclei was 0.22% for vehicle control, 0.14, 0.18 and 0.25% for DINP 1 at 500, 1000, and 2000 mg/kg bw/day, respectively, and 2.2% for cyclophosphamide at 20 mg/kg bw/day.
- Ratio of PCE/NCE (for Micronucleus assay): about 1:1 in all groups
- Statistical evaluation: No statistically significant increases in percentage of PCEs with micronuclei in DINP groups compared to vehicle control. - Conclusions:
- Interpretation of results: negative
No in vivo genotoxicity data are available on S278. No chromosome aberrations (increased frequency of micronuclei) were seen in the bone marrow erythrocytes of male mice (5/dose) given the structurally related compound, di(isononyl) phthalate (DINP 1; CAS 68515-48-0), by stomach tube on two consecutive days at up to 2000 mg/kg bw/day. - Executive summary:
No in vivo genotoxicity data are available on S278. A micronucleus test was performed to assess the potential of the structurally related compound, di(isononyl) phthalate (DINP 1; CAS 68515-48-0), to induce chromosome aberrations in the bone marrow of mice.
Briefly, young male mice (5/dose) were administered DINP 1 by stomach tube at 0, 500, 1000, or 2000 mg/kg bw/day on two consecutive days. On the third day the animals were sacrificed, and slides were prepared from bone marrow erythrocytes. A total of 1000 erythrocytes were counted from each animal, and the total numbers of polychromatic (PCE) and normochromatic (NCE) erythrocytes were tabulated. A total of 1000 PCEs were evaluated for the presence of micronuclei in the treated group, and compared to that from the vehicle control group (no further details given on identity and dose of vehicle control) and the positive control group (receiving cyclophosphamide at 20 mg/kg bw/day).
Frequency of micronucleus formation was not elevated at any dose in the treated animals compared to the vehicle controls, and there was no evidence of toxicity (examination not described) at up to 2000 mg/kg bw/day. Treatment with the positive control induced a statistically significant increase in micronuclei formation, confirming the sensitivity of the testing procedure.
In conclusion, DINP 1 was inactive in an in vivo micronucleus test in which male mice (5/dose) were administered this phthalate by stomach tube at up to 2000 mg/kg bw/day on two consecutive days, and bone marrow erythrocytes were assessed for micronuclei formation on the third day.
- Reason / purpose for cross-reference:
- reference to same study
Data source
Reference
- Reference Type:
- publication
- Title:
- Di(isononyl) phthalate (DINP) and di(isodecyl) phthalate (DIDP) are not mutagenic
- Author:
- McKee RH, Przygoda RT, Chirdon MA, Engelhardt G, Stanley M
- Year:
- 2 000
- Bibliographic source:
- J Appl Toxicol 20: 491-497
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
- Deviations:
- not specified
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)
- Deviations:
- not specified
- GLP compliance:
- not specified
- Type of assay:
- micronucleus assay
Test material
- Reference substance name:
- Di(isonyl) phthalate
- IUPAC Name:
- Di(isonyl) phthalate
- Reference substance name:
- 1,2-Benzenedicarboxylic acid, di-C8-10-branched alkyl esters, C9-rich
- EC Number:
- 271-090-9
- EC Name:
- 1,2-Benzenedicarboxylic acid, di-C8-10-branched alkyl esters, C9-rich
- Cas Number:
- 68515-48-0
- IUPAC Name:
- bis(7-methyloctyl) phthalate
- Reference substance name:
- DINP 1
- IUPAC Name:
- DINP 1
- Details on test material:
- - Name of test material (as cited in study report): DINP 1
- Molecular formula (if other than submission substance): C26H42O4
- Molecular weight (if other than submission substance): 420 [g/mol]
- Analytical purity: >99%
- Impurities (identity and concentrations): no data
- Composition of test material, percentage of components: no data
- Isomers composition: DINP 1 is produced from an alcohol feedstock that consists of roughly equivalent amounts of 3,4-, 4,6-, 3,5-, 4,5-, and 5,6-dimethylheptanol-1.
- Purity test date: no data
- Lot/batch No.: no data
- Expiration date of the lot/batch: no data
- Stability under test conditions: no data
- Storage condition of test material: no data
Constituent 1
Constituent 2
Constituent 3
Test animals
- Species:
- mouse
- Strain:
- CD-1
- Sex:
- male
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Laboratories, Portage, MI
- Age at study initiation: ca. 6-9 weeks
- Weight at study initiation: 17-35 g (although 17 g might include females used in another experiment)
- Assigned to test groups randomly: [no/yes, under following basis: ] no data
- Fasting period before study: no data
- Housing: no data
- Diet (e.g. ad libitum): no data
- Water (e.g. ad libitum): no data
- Acclimation period: no data
ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): no data
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- - Vehicle(s)/solvent(s) used: vehicle control used, but identity not described in publication
- Details on exposure:
- no data
- Duration of treatment / exposure:
- Two consecutive days
- Frequency of treatment:
- Daily
- Post exposure period:
- One day
Doses / concentrationsopen allclose all
- Remarks:
- Doses / Concentrations:
500
Basis:
actual ingested
mg/kg bw/day
- Remarks:
- Doses / Concentrations:
1000
Basis:
actual ingested
mg/kg bw/day
- Remarks:
- Doses / Concentrations:
2000
Basis:
actual ingested
mg/kg bw/day
- No. of animals per sex per dose:
- 5 males
- Control animals:
- yes, concurrent vehicle
- Positive control(s):
- cyclophosphamide
- Justification for choice of positive control(s): no data
- Route of administration: presumably gavage
- Doses / concentrations: 20 mg/kg bw/day
Examinations
- Tissues and cell types examined:
- Erythrocytes from bone marrow
- Details of tissue and slide preparation:
- CRITERIA FOR DOSE SELECTION:
No toxicity seen in a range-finding study at 2000 mg/kg bw/day.
TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
Treated on two consecutive days, and mice sacrificed on the third day.
DETAILS OF SLIDE PREPARATION:
Briefly, both femurs were removed from each treated mouse and the proximal ends were cut to expose the bone marrow, which was then aspirated with fetal bovine serum into a centrifuge tube. The cells were collected by centrifugation and slides were prepared. After fixation in methanol, the slides were stained with acridine orange for about 1-2 minutes.
METHOD OF ANALYSIS:
The slides were evaluated at 400x by fluorescence microscopy. A total of 1000 erythrocytes were counted from each animal, and the total numbers of polychromatic (PCE) and normochromatic (NCE) erythrocytes were tabulated. A total of 1000 PCEs were evaluated for the presence of micronuclei.
OTHER: - Evaluation criteria:
- no data
- Statistics:
- Statisitical analysis included means and standard deviations of the micronucleus test data, and a test of equality of means was performed by standard one-way analysis of variance.
Results and discussion
Test results
- Sex:
- male
- Genotoxicity:
- negative
- Remarks:
- at 2000 mg/kg bw/day
- Toxicity:
- no effects
- Remarks:
- at 2000 mg/kg bw/day
- Vehicle controls validity:
- not specified
- Negative controls validity:
- not examined
- Positive controls validity:
- valid
- Additional information on results:
- RESULTS OF RANGE-FINDING STUDY
- Dose range: up to 2000 mg/kg bw/day (no details given of other dose levels)
- Solubility: no data
- Clinical signs of toxicity in test animals: no evidence of toxicity (extent of examination unclear from report)
- Evidence of cytotoxicity in tissue analyzed: no data
RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): Percentage PCEs with micronuclei was 0.22% for vehicle control, 0.14, 0.18 and 0.25% for DINP 1 at 500, 1000, and 2000 mg/kg bw/day, respectively, and 2.2% for cyclophosphamide at 20 mg/kg bw/day.
- Ratio of PCE/NCE (for Micronucleus assay): about 1:1 in all groups
- Statistical evaluation: No statistically significant increases in percentage of PCEs with micronuclei in DINP groups compared to vehicle control.
Applicant's summary and conclusion
- Conclusions:
- Interpretation of results: negative
No in vivo genotoxicity data are available on S278. No chromosome aberrations (increased frequency of micronuclei) were seen in the bone marrow erythrocytes of male mice (5/dose) given the structurally related compound, di(isononyl) phthalate (DINP 1; CAS 68515-48-0), by stomach tube on two consecutive days at up to 2000 mg/kg bw/day. - Executive summary:
No in vivo genotoxicity data are available on S278. A micronucleus test was performed to assess the potential of the structurally related compound, di(isononyl) phthalate (DINP 1; CAS 68515-48-0), to induce chromosome aberrations in the bone marrow of mice.
Briefly, young male mice (5/dose) were administered DINP 1 by stomach tube at 0, 500, 1000, or 2000 mg/kg bw/day on two consecutive days. On the third day the animals were sacrificed, and slides were prepared from bone marrow erythrocytes. A total of 1000 erythrocytes were counted from each animal, and the total numbers of polychromatic (PCE) and normochromatic (NCE) erythrocytes were tabulated. A total of 1000 PCEs were evaluated for the presence of micronuclei in the treated group, and compared to that from the vehicle control group (no further details given on identity and dose of vehicle control) and the positive control group (receiving cyclophosphamide at 20 mg/kg bw/day).
Frequency of micronucleus formation was not elevated at any dose in the treated animals compared to the vehicle controls, and there was no evidence of toxicity (examination not described) at up to 2000 mg/kg bw/day. Treatment with the positive control induced a statistically significant increase in micronuclei formation, confirming the sensitivity of the testing procedure.
In conclusion, DINP 1 was inactive in an in vivo micronucleus test in which male mice (5/dose) were administered this phthalate by stomach tube at up to 2000 mg/kg bw/day on two consecutive days, and bone marrow erythrocytes were assessed for micronuclei formation on the third day.
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