Oxybispropanediol

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:

26 June - 03 August 2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was performed according to OECD and EC guidelines and according to GLP principles.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2009

Report date:

2009

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

Oxybispropanediol

EC Number:

261-605-5

EC Name:

Oxybispropanediol

Cas Number:

59113-36-9

Molecular formula:

C6H14O5

IUPAC Name:

3-(2,3-dihydroxypropoxy)propane-1,2-diol

Details on test material:

- Name of test material (as cited in study report): Diglycerol
- Substance type: liquid
- Physical state: Colourless viscous liquid
- Analytical purity:
90.7% Diglycerol
6.1% Cyclic triglycerol
2.4% triglycerol

- Purity test date: no data
- Lot/batch No.: RBA071205A
- Expiration date of the lot/batch: 31 May 2010
- Stability under test conditions: stable
- Storage condition of test material: At room temperature in the dark

Method

Species / strain

Species / strain / cell type:

lymphocytes:

Details on mammalian cell type (if applicable):

Cultured peripheral human lymphocytes

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

phenobarbital and ß-naphthoflavone induced rat liver S9-mix

Test concentrations with justification for top dose:

Dose range finding test: with 33, 100, 333, 1000 and 1662 µg diglycerol/ml culture medium with and without S9-mix.

First test: 333, 1000 and 1662 µg/ml culture medium without and with S9-mix (3 h exposure time, 24 h fixation time)

Second test: 333, 1000 and 1662 µg/ml culture medium without S9-mix (24 and 48 h exposure time, 24 and 48 h fixation time)
333, 1000 and 1662 µg/ml culture medium with S9-mix (3 h exposure time, 48 h fixation time)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: RPMI 1640 medium
- Justification for choice of solvent/vehicle: Diglycerol concentrations were used within 2.5 hours after preparation and filter (0.22 µm)-sterilized.

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium


DURATION
- Preincubation period: 48 h
- Exposure duration: 3h (with and without S9 mix), 24h or 48h (without S9 mix)
- Fixation time (start of exposure up to fixation or harvest of cells): 24h or 48h


SPINDLE INHIBITOR (cytogenetic assays): colchicine (0.5 µg/ml medium)
STAIN (for cytogenetic assays): 5% (v/v) Giemsa


NUMBER OF REPLICATIONS: duplicates in two independent experiments


NUMBER OF CELLS EVALUATED: 100 metaphase chromosome spreads per culture


DETERMINATION OF CYTOTOXICITY
- Method: mitotic index of each culture was determined by counting the number of
metaphases per 1000 cells.

- Determination of polyploidy: yes
- Determination of endoreplication: yes

Evaluation criteria:

A test substance was considered positive (clastogenic) in the chromosome aberration test if:
a) It induced a dose-related statistically significant (Chi-square test, one-sided, p < 0.05) increase in the number of cells with chromosome aberrations.
b) A statistically significant and biologically relevant increase in the frequencies of the number of cells with chromosome aberrations was observed in the absence of a clear dose-response relationship.

Statistics:

The incidence of aberrant cells (cells with one or more chromosome aberrations, gaps included or excluded) for each exposure group outside the laboratory historical control data range was compared to that of the solvent control using Chi-square statistics.

Results and discussion

Test results

Species / strain:

lymphocytes: Cultured peripheral human lymphocytes

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

First cytogenetic assay:
All dose levels were selected for scoring of chromosome aberrations. Both in the absence and presence of S9-mix, diglycerol did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations.
Both in the absence and presence of S9-mix, diglycerol did not increase the number of polyploid cells and cells with endoreduplicated chromosomes.

Second cytogenetic assay:
All dose levels were selected for scoring of chromosome aberrations. Both in the absence and presence of S9-mix, Diglycerol did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations.
Both in the absence and presence of S9-mix, Diglycerol did not increase the number of polyploid cells and cells with endoreduplicated chromosomes.

Any other information on results incl. tables

The number of cells with chromosome aberrations found in the solvent control cultures was within the laboratory historical control data range. The number of polyploid cells and cells with endoreduplicated chromosomes in the solvent control cultures was within the laboratory historical control data range. The positive control chemicals both produced statistically significant increases in the frequency of aberrant cells. It was therefore concluded that the test conditions were adequate and that the metabolic activation system (S9-mix) functioned properly.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

Both in the absence and presence of S9-mix Diglycerol did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in two independent experiments.No effects of Diglycerol on the number of polyploid cells and cells with endoreduplicated chromosomes were observed both in the absence and presence of S9-mix.

Executive summary:

The ability of Diglycerol to induce chromosome aberrations in human peripheral lymphocytes was investigated in two independent experiments with and without metabolic activation according to OECD 473 and GLP. No effects of diglycerol on the number of polyploid cells and cells with endoreduplicated chromosomes were observed both in the absence and presence of metabolic activation. Therefore it was concluded that diglycerol does not disturb mitotic processes and cell cycle progression and does not induce numerical chromosome aberrations. Diglycerol did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of metabolic activation. It was concluded that diglycerol is not clastogenic.