Isopentyl propionate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Experimental study on genetic toxicity in vitro was conducted by M. Ishidate al.( Fd Chem. Toxic.,!984) to determine the mutagenic nature of target substance Isoamy Propionate (105-68-0). This report presents data from primary screening by the Ames test on 200 food additives used in Japan for Isoamyl Propionate (105-68-0).The samples were supplied from the Japan Food Additives Association, Tokyo, at the request of the Ministry of Health and Welfare of Japan, where the purity and quality of each sample were checked. Reverse mutation assays using S. Typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 were carried out at the concentration of 0.1 – 10 mg/plate in the presence and absence of S9 .Acetone was used as a solvent since test compound is insoluble in water. All samples were kept in a refrigerator before use. Reverse mutation assays using S. Typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 were carried out according to the method of Ames, McCann & Yamasaki. All these test strains were originally provided by Dr B. N. Ames, University of California, Berkeley, USA. The liver microsome fraction (S-9) was prepared from the liver of Fischer rats (Charles River Japan Co.) pre-treated 5 days before with polychlorinated biphenyls (500 mg/kg body weight of Kanechlor KC-400 in olive oil, ip). The reaction mixture (S-9 mix) contained 5 mM-glucose 6-phosphate, 4mM-NADPH, 4mM-NADH, 33mM-KCl, 8 mM-MgCI2, 100 mM-phosphate buffer (pH 7.4) and 3.75 ml S-9 (129 mg protein) in a total volume of 12.5 ml. Cells cultured overnight were pre-incubated with both the test sample and the S-9 mix for 20 min at 37°C before plating. Duplicate plates were used for each of six different concentrations of the sample.The number of revertant (his +) colonies was scored after incubation at 37°C for 2 days. The result was considered positive if the number of colonies found was twice the number in the control (exposed to the appropriate solvent or untreated). A negative result indicates that no significant increases in the number of revertant colonies were detected in anyS. typhimuriumstrains at the maximum dose.At the maximum dose of 10mg/plate, the test chemical was non mutagenic. Therefore Isoamyl Propionate was considered to be non mutagenic in AMES test..Hence the test chemical cannot be classified as gene mutant in vitro.

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

Data is from publication.

Qualifier:

according to guideline

Guideline:

other: as per mentioned below

Principles of method if other than guideline:

Reverse mutation assays using S. typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 were carried out according to the method of Ames, McCann & Yamasaki.

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

Name of test material (as cited in study report): Isoamyl Propionate
Substance type: Organic
Physical state: Liquid
Supplied from Japan Food Additives Association, Tokyo, at the request of the Ministry of Health and Welfare of Japan.

Target gene:

Histidine

Species / strain / cell type:

S. typhimurium, other: Salmonella Typhirium TA92, TA1535, TA100, TA1537, TA94 and TA98

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

not specified

Metabolic activation:

with and without

Metabolic activation system:

S-9 was prepared from the liver of Fischer rats ( pretreated 5 days before with polychlorinated biphenyls (500 mg/kg body weight of Kanechlor KC-400 in olive oil, ip).

Test concentrations with justification for top dose:

0.1 – 10 mg/plate

Vehicle / solvent:

Solvent used: DMSO
Justification for choice of solvent/vehicle: Since the chemical was insoluble in water

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

not specified

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium; in agar (plate incorporation); preincubation; in suspension; as impregnation on paper disk
: pre incubation

DURATION
Preincubation period: Cells cultured overnight were pre-incubated with both the test sample and the S-9 mix for 20 min at 37°C before plating. The number of revertant (his +) colonies was scored after incubation at 37°C for 2 days
Exposure duration: 2 days

NUMBER OF REPLICATIONS: Duplicate plates were used for each of six different concentrations of the sample.

Rationale for test conditions:

Not specified

Evaluation criteria:

The test result was considered to be negative.if there were no significant increases in the numbers of revertant colonies detected in any S. typhimurium strains at the maximum dose.

Statistics:

Yes

Species / strain:

S. typhimurium, other: Salmonella Typhimurium TA92, TA1535, TA100, TA1537, TA94 and 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:

not specified

Additional information on results:

The liver microsome fraction (S-9) was prepared from the liver of Fischer rats (Charles River Japan Co.) pretreated 5 days before with polychlorinated biphenyls (500 mg/kg body weight of Kanechlor KC-400 in olive oil, ip). The reaction mixture (S-9 mix) contained 5 mM-glucose 6-phosphate, 4mM-NADPH, 4mM-NADH, 33mM-KC1, 8 mM-MgCI 2, 100 mM-phosphate buffer (pH 7.4) and 3.75 ml S-9 (129 mg protein) in a total volume of 12.5 ml.

Remarks on result:

other: No mutagenic effect were observed.

Conclusions:

Isoamyl Propionate (105-68-0) was evaluated for its mutagenic potential.A negative result indicates that no significant increases in the number of revertant colonies were detected in any S. typhimurium strains at the maximum dose.The test chemical was tested by the AMES test using 6 strains of S.typhimurium.At the maximum dose of 10mg/plate, the test chemical was non mutagenic

Executive summary:

This report presents data from primary screening by the Ames test on 200 food additives used in Japan for Isoamyl Propionate (105-68-0).The samples were supplied from the Japan Food Additives Association, Tokyo, at the request of the Ministry of Health and Welfare of Japan, where the purity and quality of each sample were checked.

Reverse mutation assays usingS. typhimuriumstrains TA92, TA1535, TA100, TA1537, TA94 and TA98 were carried out at the concentraton of 0.1 – 10 mg/plate in the presence and absence of S9 .Acetone was used as a solvent since test compound is insoluble in water.All samples were kept in a refrigerator before use.Reverse mutation assays usingS. typhimuriumstrains TA92, TA1535, TA100, TA1537, TA94 and TA98 were carried out according to the method of Ames, McCann & Yamasaki.All these test strains were originally provided by Dr B. N. Ames, University of California, Berkeley, USA. The liver microsome fraction (S-9) was prepared from the liver of Fischer rats (Charles River Japan Co.) pretreated 5 days before with polychlorinated biphenyls (500 mg/kg body weight of Kanechlor KC-400 in olive oil, ip). The reaction mixture (S-9 mix) contained 5 mM-glucose 6-phosphate, 4mM-NADPH, 4mM-NADH, 33mM-KCl, 8 mM-MgCI₂, 100 mM-phosphate buffer (pH 7.4) and 3.75 ml S-9 (129 mg protein) in a total volume of 12.5 ml. Cells cultured overnight were pre-incubated with both the test sample and the S-9 mix for 20 min at 37°C before plating. Duplicate plates were used for each of six different concentrations of the sample.The number of revertant (his +) colonies was scored after incubation at 37°C for 2 days. The result was considered positive if the number of colonies found was twice the number in the control (exposed to the appropriate solvent or untreated). A negative result indicates that no significant increases in the number of revertant colonies were detected in anyS. typhimuriumstrains at the maximum dose.At the maximum dose of 10mg/plate, the test chemical was non mutagenic. Therefore Isoamyl Propionate was considered to be non mutagenic in AMES test..Hence the test chemical cannot be classified as gene mutant in vitro.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Genotoxicity In-vitro

Various publications were reviewed to determine the mutagenic nature of Isoamyl Propionate (105-68-0). The studies are as mentioned below:

Experimental study on genetic toxicity in vitro was conducted by M. Ishidate al.( Fd Chem. Toxic.,!984) to determine the mutagenic nature of target substance Isoamy Propionate (105-68-0). This report presents data from primary screening by the Ames test on 200 food additives used in Japan for Isoamyl Propionate (105-68-0).The samples were supplied from the Japan Food Additives Association, Tokyo, at the request of the Ministry of Health and Welfare of Japan, where the purity and quality of each sample were checked. Reverse mutation assays using S. Typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 were carried out at the concentration of 0.1 – 10 mg/plate in the presence and absence of S9 .Acetone was used as a solvent since test compound is insoluble in water. All samples were kept in a refrigerator before use. Reverse mutation assays using S. Typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 were carried out according to the method of Ames, McCann & Yamasaki. All these test strains were originally provided by Dr B. N. Ames, University of California, Berkeley, USA. The liver microsome fraction (S-9) was prepared from the liver of Fischer rats (Charles River Japan Co.) pre-treated 5 days before with polychlorinated biphenyls (500 mg/kg body weight of Kanechlor KC-400 in olive oil, ip). The reaction mixture (S-9 mix) contained 5 mM-glucose 6-phosphate, 4mM-NADPH, 4mM-NADH, 33mM-KCl, 8 mM-MgCI2, 100 mM-phosphate buffer (pH 7.4) and 3.75 ml S-9 (129 mg protein) in a total volume of 12.5 ml. Cells cultured overnight were pre-incubated with both the test sample and the S-9 mix for 20 min at 37°C before plating. Duplicate plates were used for each of six different concentrations of the sample.The number of revertant (his +) colonies was scored after incubation at 37°C for 2 days. The result was considered positive if the number of colonies found was twice the number in the control (exposed to the appropriate solvent or untreated). A negative result indicates that no significant increases in the number of revertant colonies were detected in anyS. typhimuriumstrains at the maximum dose.At the maximum dose of 10mg/plate, the test chemical was non mutagenic. Therefore Isoamyl Propionate was considered to be non mutagenic in AMES test..Hence the test chemical cannot be classified as gene mutant in vitro.

 

Supported by experimental study on mammalian cell line,which is performed by M. Ishidate al. (Food Chem. Toxic., 1984) to determine the mutagenic nature of Isoamy Propionate (105-68-0). Chromosomal aberration tests in vitro using a Chinese hamster fibroblast cell line were carried out on Isoamyl Propionate (105-68-0).The maximum dose of each sample was selected by a preliminary test in which the dose needed for 50% cell-growth inhibition was estimated using a cell densitometer.The cell line was originally established from the lung of a newborn female at the Cancer Research Institute, Tokyo was maintained by 4-day passages in Minimum Essential Medium (MEM; GIBCO) supplemented by 10% calf serum. The modal chromosome number is 25 and the doubling time was approximately 15 hr. Sodium carboxymethyl cellulose was used as solvent for the test chemical, since it is insoluble in water. Chromosome preparations were made as follows: Colcemid (final concentration 0.2µg/ml) was added to the culture 2 hr before cell harvesting. The cells were then trypsinized and suspended in a hypotonic KCI solution (0.075 M) for 13 min at room temperature. After centrifugation the cells were fixed with acetic acid-methanol (1:3, v/v) and spread on clean glass slides. After air-drying, the slides were stained with Geimsa solution (1.5%, at pH 6.8) for 12-15 min. A hundred well-spread metaphases were observed under the microscope (x 600 with a no cover objective lens). The incidence of polyploid cells as well as of cells with structural chromosomal aberrations such as chromatid or chromosome gaps, breaks, exchanges, ring formations, fragmentations and others, was recorded on each culture plate.Untreated cells and solvent-treated cells served as negative controls, in which the incidence of aberrations was usually less than 3.0%. The results were considered to be negative if the incidence was less than 4.9%, equivocal if it was between 5.0 and 9.9%, and positive if it was more than 10.0%. When no reasonable dose-response relationships were found, additional experiments were carried out at similar dose levels. For a quantitative evaluation of the clastogenic potential of the positive samples, the D20 was calculated, which is the dose (mg/ml) at which structural aberrations (including gaps) were detected in 20% of the metaphases observed. In addition, the TR value was calculated, which indicates the frequency of cells with exchange-type aberrations per unit dose (mg/ml).Since the incidence of aberrations for the test chemical was 2.0% at the maximum dose, it can be concluded that the test chemical is non mutagenic. Hence the test substance cannot be classified as gene mutant in vitro.

Another supporting study conducted by M. Ishidate et. al (Mutation Research,1988) to determine the mutagenic nature of Isoamy Propionate (105-68-0). A chromosomal aberration test on Isoamyl Propionate (105-68-0) using mammalian cells in culture was performed. Chinese hamster lung fibroblast cell lines were used for the test. A clonal sub-line of fibroblasts derived from lung tissue (Koyama et al, 1970).The cell line was exposed to 2000µg/ml of test chemical for 48 hours without metabolic activation. No mutagenic or clastogenic activity was observed for the test chemical. Therefore Isoamyl Propionate was considered to be non mutagenic in mammalian cell line. Hence the test substance cannot be classified as gene mutant in vitro.

Further supported by another experimental study conducted by Kunita n (OSAKA-FURITSU KOSHU EISEI KENKYU HOKOKU SHOKUHIN EISEI HEN, 1978) to determine the mutagenic nature of Isoamy Propionate (105-68-0 . The mutagenic properties of 3-methylbutyl propionate (105-68-0) were evaluated using BACILLUS SUBTILIS, H17 (rec +) and M45 (rec -) REC assay. The maximum dose of test chemical evaluated was 17µg/disk. The test was conducted in the absence of metabolic activation. The test chemical was found to be non mutagenic in the REC assay. Therefore 3-methylbutyl propionate was considered to be non mutagenic in REC assay. Hence the test substance cannot be classified as gene mutant in vitro.

Based on the data available for the target chemical, it is concluded that Isoamyl Propionate (105-68-0) does not exhibit gene mutation in vitro. Hence the test chemical is not likely to classify as a gene mutant in vitro.

Justification for classification or non-classification

Thus based on the above annotation for the target chemical, it is concluded that Isoamyl Propionate (105-68-0) does not exhibit gene mutation in vitro. Hence the test chemical is not likely to classify as a gene mutant in vitro.