Benzyl butyl phthalate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

no data

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: NTP study carried out to national standards that applied at the time.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

EPA OPPTS 870.5375 - In vitro Mammalian Chromosome Aberration Test

Deviations:

yes

Remarks:

Only one culture per dose; two independent cultures per dose are recommended

GLP compliance:

not specified

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

Benzyl butyl phthalate

EC Number:

201-622-7

EC Name:

Benzyl butyl phthalate

Cas Number:

85-68-7

Molecular formula:

C19H20O4

IUPAC Name:

1-benzyl 2-butyl benzene-1,2-dicarboxylate

Details on test material:

- Name of test material (as cited in study report): butyl benzyl phthalate
- Physical state: solid
- Analytical purity: no data
- Impurities (identity and concentrations): no data
- 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

Method

Target gene:

not applicable

Species / strain

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

- Type and identity of media: McCoy's 5a medium supplemented with 10% fetal calf serum, L-glutamine and antibiotics
- Properly maintained: no data
- Periodically checked for Mycoplasma contamination: no data
- Periodically checked for karyotype stability: no data
- Periodically "cleansed" against high spontaneous background: no data

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

S9 fraction prepared from Aroclor 1254-induced rat liver

Test concentrations with justification for top dose:

0, 125, 400 or 1250 ug/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: included in list of recommended solvents

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

triethylenemelamine

Remarks:

Migrated to IUCLID6: 0.15 ug/mL without S9

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

Migrated to IUCLID6: 15.0 ug/mL with S9

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium


DURATION
- Exposure duration: Without S9 the cells were incubated with the test substance for 12 hours, then colcemid added and incubated for a further 2 hr. With S9 the cells were exposed to the test substance for 2 hr, then incubated for a further 12 hr in fresh medium. Colcemid was added for the final 2 hr of incubation

- Fixation time (start of exposure up to fixation or harvest of cells): 14 hr


SPINDLE INHIBITOR (cytogenetic assays): colcemid

STAIN (for cytogenetic assays): Giemsa


NUMBER OF REPLICATIONS: one


NUMBER OF CELLS EVALUATED: 100 cells/dose


DETERMINATION OF CYTOTOXICITY
- Method: other: doses based on the cytotoxicity seen in a sister chromatid exchange assay in the same cell type.


OTHER EXAMINATIONS:
- Determination of polyploidy: no
- Determination of endoreplication: yes

Evaluation criteria:

Only cells containing 21 +/- 2 chromosomes were scored; all types of aberrations were included. Gaps and endoreduplications were recorded but not included in the totals.

Statistics:

Linear regression analysis of the percentage of cells with aberrations versus the log-dose of the dose was used to test for trend.

Results and discussion

Test results

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
no data


RANGE-FINDING/SCREENING STUDIES: based on a previous assay for sister chromatid exchanges using the same cell type

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

Butyl benzyl phthalate did not induce chromosome aberrations when tested in an in vitro assay with Chinese hamster ovary cells, both in the presence and absence of a rat liver metabolic activation fraction.

Executive summary:

Butyl benzyl phthalate was assessed for its ability to induce chromosome aberrations in Chinese hamster ovary cells.

Cells were incubated with concentrations of the test substance of up to 1250 ug/mL for 2 hr with S9, or 14 hr without S9. Cells that had been exposed with S9 were placed in fresh medium and incubated for a further 12 hr.; colcemid was added as a spindle inhibitor to all cultures 2 hr before harvesting the cells.After fixation and staining, 100 first-division metaphases per dose were scored for all types of chromosome aberrations.

The test substance did not increase the frequency of aberrations, compared to the vehicle controls.

Butyl benzyl phthalate did not induce chromosome aberrations in an in vitro assay with Chinese hamster ovary cells, either in the presence or absence of a rat liver metabolic activation fraction.