4-chlorobenzophenone

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

data is from peer reviewed journals

Data source

Reference

Reference Type:

publication

Title:

Modified AMES assay for various test chemicals

Author:

Rexroat et.al

Year:

1995

Bibliographic source:

Mutation Research, 1995

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

other: OECD 471

Principles of method if other than guideline:

Ames Assay was performed to determine the mutagenic potential of the test chemical

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

2,4'-dichlorobenzophenone

EC Number:

201-596-7

EC Name:

2,4'-dichlorobenzophenone

Cas Number:

85-29-0

Molecular formula:

C13H8Cl2O

IUPAC Name:

Benzophenone, 2,4'-dichloro

Test material form:

solid

Details on test material:

Name of the test chemical: 2,4'-dichlorobenzophenone
IUPAC name: (2-chlorophenyl)(4-chlorophenyl)methanone
Molecular Weight: 251.1 g/mol
Molecular Formula: C13H8Cl2O
Substance Type: Organic
Physical State: solid

Method

Target gene:

Salmonella -Histidine
tryptophan - E.Coli

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium, other: S. typhimurium LT-2: TA1535, TA1537, TA100, and TA98

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Cytokinesis block (if used):

no data available

Metabolic activation:

with and without

Metabolic activation system:

The test was also conducted with metabolic activation using a rat post-mitochondrial fraction (Aroclor 1254-induced S9) which was prepared by Lilly Research Laboratories or obtained from a commercial source (Molecular Toxicology, Annapolis, Maryland). The S9 was validated with appropriate positive controls prior to use

Test concentrations with justification for top dose:

100 to 1000 microgram/ml.

Vehicle / solvent:

solvent used name not mentioned

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

name not mentioned

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

2-acetylaminofluorene

9-aminoacridine

2-nitrofluorene

Details on test system and experimental conditions:

NUMBER OF REPLICATIONS:
- Number of cultures per concentration (single, duplicate, triplicate) : triplicates
- Number of independent experiments : single

METHOD OF TREATMENT/ EXPOSURE:
- Cell density at seeding (if applicable):
- Test substance added in medium; in agar (plate incorporation); preincubation; in suspension; as impregnation on paper disk : agar medium

TREATMENT AND HARVEST SCHEDULE:
- Preincubation period, if applicable: 48 h
- Exposure duration/duration of treatment: 48h
- Harvest time after the end of treatment (sampling/recovery times): no data available


- OTHER: Five bacterial strains were used in both the Ames assay.They include four auxotrophs of S. typhimurium LT-2: TA1535, TA1537, TA100, and TA98 and a tryptophan auxotroph derived from E. coli: WP2uvrA . Salmonella typhimurium strains were obtained from Dr. Bruce N. Ames, and the E. coli strain was obtained from Dr. Brian A. Bridges of the MRC Cell Mutation Unit, Sussex, UK. The genotypes of these strains were confirmed using the procedure of Maron and Ames (1983). The overnight cultures were prepared from frozen stock (-150°C) by inoculating 0.2 ml of each tester strain in 20 ml of 2.5%
Oxoid Nutrient Broth No. 2 (Oxoid LTD). Minimal agar medium included 100 ml of 20% glucose; 80 ml of 10 x histidine-tryptophan-biotin(each his-tryp-bio, 0.5 mM); 40 ml of 25 × Vogel- Bonner salts (VB); and 280 ml Milli-Q water. The above mixture was warmed to 50°C and QS to 1 liter with melted 2.5% Difco Bacto agar. Final concentrations were 2% glucose, 1 × VB salts, and 0.04 mM of histryp-bio in 1.25% agar medium. A stock concentration of the test chemical was prepared at 100 mg/ml, and three 1:10 serial dilutions were performed. A concentration of 1000 microgram/ml of test compoundwas prepared by adding 10 ml of minimal agar to 0.1 ml of a 100 mg/ml solution of compound in the appropriate vehicle. The top agar was thoroughly mixed and poured onto the appropriate base layer plate. This resulted in a ten-fold concentration gradient across the plate ranging from 100 to 1000 microgram/ml.

Rationale for test conditions:

no data available

Evaluation criteria:

A test article was considered to have induced a positive response for bacterial mutation when a concentration-related increase in revertants was observed in which the number of revertants exceeded
the value of the vehicle control by at least two-fold (strains TA98, TA100, and WP2uvrA-) or a least three-fold (strains TA1535 and TA1537), for two successive concentrations of the test article.

Statistics:

no data available

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium, other: S. typhimurium LT-2: TA1535, TA1537, TA100, and TA98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not specified

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

no data available

Remarks on result:

other: not mutagenic

Applicant's summary and conclusion

Conclusions:

The test chemical failed to induce genetic toxicity when tested on the Salmonella typhimurium LT-2: TA1535, TA1537, TA100, and TA98 and a tryptophan auxotroph derived from E. coli: WP2uvrA strains in the presence and absence of S9 metabolic activation system. Hence, the test chemical can be considered to be non-genotoxic in nature.

Executive summary:

Ames Assay was performed to determine the mutagenic potential of the test chemical.Five bacterial strains were used in the Ames assay. They include four auxotrophs of Salmonella typhimurium LT-2: TA1535, TA1537, TA100, and TA98 and a tryptophan auxotroph derived from E. coli: WP2uvrA.Salmonella typhimurium strains were obtained from Dr. Bruce N. Ames, and the E. coli strain was obtained from Dr. Brian A. Bridges of the MRC Cell Mutation Unit, Sussex, UK. The genotypes of these strains were confirmed using the procedure of Maron and Ames (1983). The overnight cultures were prepared from frozen stock (-150°C) by inoculating 0.2 ml of each tester strain in 20 ml of 2.5% Oxoid Nutrient Broth No. 2 (Oxoid LTD). Minimal agar medium included 100 ml of 20% glucose; 80 ml of 10 x histidine-tryptophan-biotin(each his-tryp-bio, 0.5 mM); 40 ml of 25 × Vogel- Bonner salts (VB); and 280 ml Milli-Q water. The above mixture was warmed to 50°C and QS to 1 liter with melted 2.5% Difco Bacto agar. Final concentrations were 2% glucose, 1 × VB salts, and 0.04 mM of histryp-bio in 1.25% agar medium. A stock concentration of the test chemical was prepared at 100 mg/ml, and three 1:10 serial dilutions were performed. A concentration of 1000 microgram/ml of test compound was prepared by adding 10 ml of minimal agar to 0.1 ml of a 100 mg/ml solution of compound in the appropriate vehicle. The top agar was thoroughly mixed and poured onto the appropriate base layer plate. This resulted in a ten-fold concentration gradient across the plate ranging from 100 to 1000 microgram/ml. The test was also conducted with metabolic activation using a rat post-mitochondrial fraction (Aroclor 1254-induced S9) which was prepared by Lilly Research Laboratories or obtained from a commercial source (Molecular Toxicology, Annapolis, Maryland). The S9 was validated with appropriate positive controls prior to use.In the Ames assay, 5 bacterial strains, 5 test concentrations, 1 solvent control, and 1 concentration per positive control were tested in triplicate, with and without metabolic activation (210 plates). Revertant colonies were counted using an Artek 880 Automated Colony Counter. A test article was considered to have induced a positive response for bacterial mutation when a concentration- related increase in revertants was observed in which the number of revertants exceeded the value of the vehicle control by at least two-fold (strains TA98, TA100, and WP2uvrA-) or a least three-fold (strains TA1535 and TA1537), for two successive concentrations of the test article. The test chemical failed to induce genetic toxicity when tested on the Salmonella typhimurium LT-2: TA1535, TA1537, TA100, and TA98 and a tryptophan auxotroph derived from E. coli: WP2uvrA strains in the presence and absence of S9 metabolic activation system. Hence, the test chemical can be considered to be non-genotoxic in nature.