Heptanoyl chloride

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:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Heptanoyl chloride

EC Number:

219-775-3

EC Name:

Heptanoyl chloride

Cas Number:

2528-61-2

Molecular formula:

C7H13ClO

IUPAC Name:

heptanoyl chloride

Details on test material:

Name of the test substance used in the study report: n-Heptansaeurechlorid
Degree of purity: 98.6%

Method

Target gene:

S. typhimurium and E. coli

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

Aroclor induced rat liver S-9 mix

Test concentrations with justification for top dose:

20 ug - 5000 ug/plate (standard plate test - all tester strains)
50 ug - 800 ug/plate (preincubation test - Salmonella strains)
62.5 ug - 1000 ug/plate (preincution test - E. coli)

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

other: with S-9 mix: 2-aminoanthracene; without S-9 mix: N-methyl-N'-nitro-N-nitrosoguanidine, 4-nitro-o-phenylendiamine, 9-aminoacridine (AAC) chloride monohydrate, N-ethyl-N'-nitro-N-nitrosoguanidin

Details on test system and experimental conditions:

Standard plate test:
Salmonella typhimurium:
The experimental procedure is based on the method of Ames et al .
Test tubes containing 2 ml portions of soft agar which consists of 100 ml agar (0.6% agar + 0.6% NaCl) and 10 ml amino acid solution (minimal amino acid solution for the determination of mutants : 0.5 mM histidine + 0.5 mM biotin) are kept in a water bath at 45°C, and the remaining components are added in the following order :
0.1 ml test solution or vehicle; 0.1 ml bacterial suspension; 0.5 ml S-9 mix (in tests with metabolic activation) or 0.5 ml phosphate buffer (in tests without metabolic activation)
After mixing, the samples are poured onto Vogel-Bonner agar plates (minimal glucose agar plates) within approx . 30 seconds .
Composition of the minimal glucose agar :
980 ml aqua dest .; 20 ml Vogel-Bonner E medium; 15 g Difco bacto agar; 20 g D-glucose, monohydrate .
After incubation at 37°C for 48 - 72 hours in the dark, the bacterial colonies (his+ revertants) are counted.

Escherichia coli:
The experimental procedure is based on the method of Ames et al .
Test tubes containing 2 ml portions of soft agar which consists of 100 ml agar (0.6% agar + 0.6% NaCl) and 10 ml amino acid solution (minimal amino acid solution for the determination of mutants : 0.5 mM tryptophan) are kept in a water bath at 45°C and the remaining components are added in the following order :
0.1 ml test solution or vehicle; 0.1 ml bacterial suspension; 0.5 ml S-9 mix (in tests with metabolic activation) or 0.5 ml phosphate buffer (in tests without metabolic activation).
After mixing, the samples are poured onto minimal agar plates within approx . 30 seconds .
Composition of the minimal agar :
The composition of the minimal agar (SA1 selective agar) is based on the description of Green, M .H .L . and Muriel, W .J , with the exception of solution E (tryptophan solution), which has been added to the soft agar before :
300 ml solution B (agar ); 100 ml solution A (saline solution); 8 ml solution C (glucose solution); 10 ml solution D (casein solution)
After incubation at 37°C for 48 - 72 hours in the dark, the bacterial colonies (trp+ revertants) are counted.


Preincubation test:
The experimental procedure is based on the method described by Yahagi et al . and Matsushima et al.
0.1 ml test solution or vehicle, 0.1 ml bacterial suspension and 0.5 ml S-9 mix are incubated at 37°C for the duration of 20 minutes . Subsequently, 2 ml of soft agar is added and, after mixing, the samples are poured onto the agar plates within approx. 30 seconds.
After incubation at 37°C for 48 - 72 hours in the dark, the bacterial colonies are counted.

Evaluation criteria:

In general, a substance to be characterized as positive in the bacterial tests has to fulfill the following requirements:
- doubling of the spontaneous mutation rate (control)
- dose-response relationship
- reproducibility of the results

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Solubility: No precipitation of the test substance was found.
Toxicity: A bacteriotoxic effect (reduced his' or trp' background growth, decrease in the number of his+ or trp+ revertants) was observed in the standard plate test from about 500 pg - 750 μg/plate onward (Salmonella strains) or at doses z 1,000 (E . coli) .
In the preincubation test bacteriotoxicity was found at doses >= 200 pg/plate (Salmonella strains) or at doses >= 500/plate (E . coli) .
Mutagenicity: An increase in the number of his+ or trp+ revertants was not observed both in the standard plate test and in the preincubation test either without S-9 mix or after the addition of a metabolizing system.

Applicant's summary and conclusion

Conclusions:

According to the results of the present study, the test substance is not mutagenic in the Ames test and in the Escherichia coli reverse mutation assay under the experimental conditions chosen here .