5-Isobenzofurancarboxylic acid, 1,3-dihydro-1,3-dioxo-, reaction products with 1-nonanol

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Deviations:

not specified

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

Trioctyl benzene-1,2,4-tricarboxylate

EC Number:

201-877-4

EC Name:

Trioctyl benzene-1,2,4-tricarboxylate

Cas Number:

89-04-3

Molecular formula:

C33H54O6

IUPAC Name:

trioctyl benzene-1,2,4-tricarboxylate

Test material form:

liquid

Specific details on test material used for the study:

- Lot/batch No.: C-120

Method

Target gene:

N/A

Species / strain

Species / strain / cell type:

other: Chinese hamster lung cells CHL/IU

Details on mammalian cell type (if applicable):

- Type and identity of media: Inactivated calf serum in Eagle MEM
- Properly maintained: No data
- Periodically checked for Mycoplasma contamination: No data
- Periodically checked for karyotype stability: No data
- Periodically "cleansed" against high spontaneous background: yes

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S9 fraction (mixed function induction of rat liver)

Test concentrations with justification for top dose:

-S9 (continuous treatment): 0, 1.3, 2.5, 5.0 mg/ml
-S9 (short term treatment): 0, 1.3, 2.5, 5.0 mg/ml
+S9 (short term treatment): 0, 1.3, 2.5, 5.0 mg/ml

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone; supplied by Wako Pure Chemical Industries Co.
- Justification for choice of solvent/vehicle: Substance is poorly soluble in water; very soluble in acetone & DMSO

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

-S9; 6 & 24 hours of exposure

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Remarks:

+S9; 6 hours exposure

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium
- Preincubation period:
- Exposure duration: For continuous treatment, cells were treated for 24 hours in the absence of S9; for short-term treatment cells were treated for 6 hours with & without S9 and cultivated with fresh media for 18 hours

SPINDLE INHIBITOR (cytogenetic assays): Colcimid 0.1 ug/mL
STAIN (for cytogenetic assays): 3% Giemsa solution

NUMBER OF REPLICATIONS: 2

NUMBER OF CELLS EVALUATED: 100 metaphases/slide of each replicate

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No cytotoxicity up to and including 5.0 mg/mL when inhibition of cell growth determined.

OTHER EXAMINATIONS:
- Determination of polyploidy:Negative
- Determination of endoreplication: No data or negative if equivalent to clastogenicity

Evaluation criteria:

A dose related increase or a reproducible increase in the number of cells with chromosome aberrations.
An increase in the number of polyploid cells.
An increase in the number of cells with endoreduplicated chromosomes

Statistics:

No data, thought to be Fisher's Exact Test

Results and discussion

Test results

Species / strain:

other: Chinese hamster lung cells (CHL/IU)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

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

Remarks:

5 mg/mL

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No
- Effects of osmolality: No
- Evaporation from medium: No
- Water solubility: poor
- Precipitation: No
- Other confounding effects: None

RANGE-FINDING/SCREENING STUDIES: yes; growth inhibition test (range of concentrations: 0.0-5.0 mg/mL

COMPARISON WITH HISTORICAL CONTROL DATA: No data

ADDITIONAL INFORMATION ON CYTOTOXICITY: No toxicity observed

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1 - Chromosome analysis of cells following short-term treatment with and without S9 mix

 

Group	Concentration (μg/mL)	S9	Exposure Time (hours)	Cell	No. of cells analysed	No. of structural aberrations							Number of cells with aberrations (%)	Polyploidy
		mix		growth index		gap	ctb	cte	csb	cse	frg	Total		(%)
Solvent control	0	-	6	100	200	0	0	0	1	0	0	1	1 (0.5)	0.0
	1250	-	6	107	200	0	1	0	0	0	0	1	1 (0.5)	0.0
Test substance	2500	-	6	111	200	0	0	0	1	0	0	1	1 (0.5)	0.0
	5000	-	6	107	200	0	1	0	2	0	0	3	3 (1.5)	0.0
MMC	0.1	-	6	113	200	4	17	76	1	2	0	96	91 (45.5)	0.0
Solvent control	0	+	6	100	200	1	1	1	2	0	0	4	3 (1.5)	0.0
TOTM	1250	+	6	105	200	0	2	0	1	2	0	5	5 (2.5)	0.0
T0TM	2500	+	6	103	200	0	0	0	1	1	0	2	2 (1.0)	0.0
TOTM	5000	+	6	94	200	0	1	1	0	0	0	2	2 (1.0)	0.0
BP	20	+	6	133	200	2	14	144	0	2	0	160	147 (73.5)	0.0

Abbreviations:

gap: chromatid gap and chromosome gap, ctb: chromatid break, cte: chromatid exchange, csb: chromosome break, cse: chromosome exchange (dicentric and ring etc.),frg: fragment,MMC: mitomycin C,BP: benzo(a)pyrene

 

 

Table 2 - Chromosome analysis of cells following continuous treatment without S9 mix

  

Group	Concentration (μg/mL)	S9	Exposure Time (hours)	Cell	No. of cells analysed	No. of structural aberrations							Number of cells with aberrations (%)	Polyploidy
		mix		growth index		gap	ctb	cte	csb	cse	frg	Total		(%)
Solvent control	0	-	24	100	200	1	0	1	1	0	0	2	2 (1.0)	0.0
	1250	-	24	98	200	1	0	0	0	0	0	0	0 (0.0)	0.0
Test substance	2500	-	24	104	200	0	0	1	1	0	0	2	2 (1.0)	0.0
	5000	-	24	107	200	2	1	0	0	0	0	1	1 (0.5)	0.0
MMC	0.03	-	24	103	200	3	20	38	2	2	0	62	61 (30.5)	0.0

 

Abbreviations:

gap: chromatid gap and chromosome gap, ctb: chromatid break, cte: chromatid exchange, csb: chromosome break, cse: chromosome exchange (dicentric and ring etc.),frg: fragment,MMC: mitomycin C

Applicant's summary and conclusion

Conclusions:

Interpretation of results: negative

In the conditions in which this study was performed the tested substance, at doses of up to 5.0 mg/mL, did not induce chromosome aberrations with or without S9 in cultured mammalian somatic cells.

Executive summary:

The substance has been assayed for its ability to cause chromosomal damage in cultured mammalian cells following in-vitro treatment in the absence and presence of S9 metabolic activation.

The substance did not induce chromosomal aberrations in human lymphocytes after in-vitro treatment