Benzimidazole

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

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 publication

Data source

Reference

Reference Type:

publication

Title:

Gene mutation toxicity study of the test chemical

Author:

Zeiger et al

Year:

1992

Bibliographic source:

Environmental and Molecular Mutagenesis

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

Ames Salmonella typhimurium test was conducted on Salmonella typhimurium strains TA100,TA1535,TA1537,TA97,TA98 to evaluate the mutagenic effect 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:

Benzimidazole

EC Number:

200-081-4

EC Name:

Benzimidazole

Cas Number:

51-17-2

Molecular formula:

C7H6N2

IUPAC Name:

1H-1,3-benzodiazole

Details on test material:

- Name of test material: Benzimidazole
- IUPAC name: 1H-benzimidazole
- Molecular formula: C7H6N2
- Molecular weight: 118.1384 g/mol
- Substance type: Organic
- Physical state: Solid
- Purity: Analyzed purity 99%+, Vendor's purity: 98%
- Impurities (identity and concentrations): No data

Method

Target gene:

Histidine

Species / strain

Species / strain / cell type:

S. typhimurium, other: TA100,TA1535,TA1537,TA97,TA98

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

with and without

Metabolic activation system:

The S-9 (9,000 x g supernatant) fractions of Aroclor 1254-induced, male Sprague-Dawley rat and male Syrian hamster livers were used

Test concentrations with justification for top dose:

0, 33, 100, 333, 1000, 3333 or 6666 µg/plate

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

9-aminoacridine

sodium azide

other: 4-nitro-o-phenylenediamine (TA98 and TA1538; Without S9); 2-aminoanthracene (All strains; with S9)

Details on test system and experimental conditions:

METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period:20 min
- Exposure duration: 2 days (48 hrs)
- Expression time (cells in growth medium): 2 days (48 hrs)
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells):no data

SELECTION AGENT (mutation assays): No data
SPINDLE INHIBITOR (cytogenetic assays):No data
STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: No data

NUMBER OF CELLS EVALUATED: No data

DETERMINATION OF CYTOTOXICITY No data
- Method: mitotic index; cloning efficiency; relative total growth; other: No data

OTHER EXAMINATIONS:
- Determination of polyploidy: No data
- Determination of endoreplication: No data
- Other: No data

OTHER: No data

Rationale for test conditions:

No data

Evaluation criteria:

Magnitude of the dose dependent increase in his+ revertants, and the shape of the dose response.

Evaluations were made at both the individual trial and chemical levels. Individual trials were judged mutagenic (+), weakly mutagenic (+ W), questionable (?),
or nonmutagenic (-), depending on the magnitude of the increase in his+ revertants, and the shape of the dose response. A trial was considered questionable (?) if the dose-response was judged insufficiently high to support a call of “+ W”, if only a single dose was elevated over the control, or if a weak increase was not dose-related.

A chemical was judged mutagenic (+) or weakly mutagenic (+W) if it produced a reproducible, dose-related response over the solvent control, under a single metabolic activation condition, in replicate trials. A chemical was judged questionable (?) if the results of individual trials were not reproducible, if increases in his+ revertants did not meet the criteria for a “+W” response, or if only single doses produced increases in hisf revertants in repeat trials. Chemicals were judged nonmutagenic (-) if they did not meet the criteria for a mutagenic or questionable response.

Statistics:

No data

Results and discussion

Test results

Species / strain:

S. typhimurium, other: TA100, TA1535, TA1537, TA97, TA98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No data
- Effects of osmolality: No data
- Evaporation from medium: No data
- Water solubility: No data
- Precipitation: No data
- Definition of acceptable cells for analysis: No data
- Other confounding effects: No data

RANGE-FINDING/SCREENING STUDIES: Benzimmidazole was run initially in a toxicity assay to determine the appropriate dose range for the mutagenicity assay. The toxicity assay was performed using TA100or the system developed by Waleh et al. [1982]. Toxic concentrations were defined as those that produced a decrease in the number of his+ colonies, or a clearing in the density of the background lawn, or both.

CYTOKINESIS BLOCK (if used)
- Distribution of mono-, bi- and multi-nucleated cells: No data

NUMBER OF CELLS WITH MICRONUCLEI
- Number of cells for each treated and control culture: No data
- Indication whether binucleate or mononucleate where appropriate: No data

HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Positive historical control data: No data
- Negative (solvent/vehicle) historical control data: No data

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Measurement of cytotoxicity used: No data
- Other observations when applicable: No data

Remarks on result:

other: No mutagenic potential

Any other information on results incl. tables

For strain TA100:

Dose	NA		10%HLI		30% HLI		10%RLI		30%RLI
µG/plate	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM
0	95	8.4	136	24.4	133	3.8	90	5.8	123	11.5
33	90	4.6
100	90	5.2	142	5.7	124	1.2	84	0.3	123	9.0
333	102	4.7	153	3.5	127	13.6	101	6.2	127	2.2
1000	110	13.9	146	4.4	125	9.2	96	4.7	114	12
3333	101	1.7	130	3.3	110	6.9	101	5.5	110	18.6
6666			104	15.7	92	2.9	87	3.7	73	3.7
POS	497	38.9	458	84.4	380	14.6	339	69.0	413	11.3

 

For strain TA1535:-

Dose	NA		10%HLI		30% HLI		10%RLI		30%RLI
µG/plate	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM
0	12	1.9	12	0.3	10	2.7	7	1.8	12	2.5
33	19	5.0
100	19	2.8	12	2	12	1.5	10	0	10	1.9
333	19	0.6	7	2	9	2.0	13	2.6	11	1.3
1000	17	1.5	9	2.7	8	0.7	9	0.7	11	2.1
3333	11	2.5	11	2.1	5	0.9	9	1.0	8	1.2
6666			5	0.9	5	1.3	7	1.8	3	1.5
POS	332	14.2	168	20.4	310	24.2	198	10.7	100	0.9

 

For strain TA1537:-

Dose	NA		10%HLI		30% HLI		10%RLI		30%RLI
µG/plate	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM
0	5	1.2	9	2.3	8	2.3	8	1.7	8	0.9
33	8	2.0
100	6	0.7	6	1.0	9	1.2	9	0.9	7	0.6
333	6	0.9	6	0.9	6	1.5	6	1.3	8	1.7
1000	8	1.5	10	2.1	6	1.9	7	2.1	9	1.7
3333	7	1.5	6	1.5	7	0.9	10	1.5	7	2.0
6666			5	2	6	0.3	5	1.7	5	2.0
POS	376	62.4	50	2.9	49	2.6	43	2.8	56	4.2

 

For strain TA97:-

Dose	NA		10%HLI		30% HLI		10%RLI		30%RLI
µG/plate	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM
0	164	12.2	166	6.1	152	7.5	177	10.7	183	9.2
33	165	6.1		12.8
100	180	4.4	174	9.8	179	5.5	191	5.2	95	6.4
333	183	9	184	2.8	178	8.2	180	15.7	190	11.7
1000	156	31.4	184	10.6	159	17.6	201	15.5	206	1.5
3333	103	3.3	171	4.0	118	2	167	16.4	99	27.0
6666			142		57	8.3	87	3.6	139	13.7
POS	986	88.5	613	20.5	396	6.8	498	3.9	440	6.9

 

 

 

 

 

 

 

 

 

For strain TA98:-

Dose	NA		10%HLI		30% HLI		10%RLI		30%RLI
µG/plate	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM	Mean	SEM
0	17	2.9	31	2.7	50	2.1	17	1.2	46	3.6
33	17	1.7
100	19	1.9	28	1.2	49	4.4	22	2.2	52	2.0
333	18	2.1	30	1.5	50	3.2	24	0.6	47	4.4
1000	17	1.9	25	1.5	49	3.3	24	4.5	53	2.3
3333	8	1.9	26	2.4	42	0.9	20	1.3	46	3.1
6666			28	1.2	32	9.1	17	3.3	31	11.7
POS	675	40.9	384	45.2	211	7.1	318	33.8	311	6.4

Applicant's summary and conclusion

Conclusions:

Non-mutagenic effects were known in Ames assay by using Salmonella typhimurium of strain TA100, TA1535, TA1537, TA97, TA98 when exposed with the test chemical in preincubation method and hence it is not likely to classify as a gene mutant in vitro.

Executive summary:

Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical. Ames assay was performed by the preincubation protocol using Salmonella typhimurium strains  TA100, TA1535, TA1537, TA97, TA98 both in the presence and absence of S9 metabolic activation system. The study was performed at dose levels of 0, 33, 100, 333, 1000, 3333 or 6666 µg/plate following a preincubation time of 20 mins. Concurrent solvent and positive controls were included in the study. The evaluation was done considering the magnitude of the dose dependent increase in his+ revertants, and the shape of the dose response. The test chemical did not cause a dose dependent increase in the number of histidine revertants in Salmonella typhimurium strains  TA100, TA1535, TA1537, TA97, TA98 both with and witthout S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.