polymeric zinc propylenebis(dithiocarbamate);propineb (ISO)

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1987/06/30 to 1987/10/22

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1988

Report date:

1988

Materials and methods

Test guideline

Qualifier:

no guideline available

Principles of method if other than guideline:

The study conforms to the OECD Principles of Good Laboratory Practice (Bundesanzeiger Nr. 42a/7of, 2nd of March 1983). The study is not fully compliant with the current guideline with respect to the number of cells exposed and background mutation rate, and is therefore of limited sensitivity.

GLP compliance:

yes (incl. QA statement)

Type of assay:

in vitro mammalian cell gene mutation test using the Hprt and xprt genes

Test material

Test material information

Constituent 1

Reference substance name:

polymeric zinc 1,2-propylenebis(dithiocarbamate)

EC Number:

620-365-5

Cas Number:

9016-72-2

IUPAC Name:

polymeric zinc 1,2-propylenebis(dithiocarbamate)

Test material form:

solid

Remarks:

Powder

Method

Target gene:

HGPRT locus

Species / strain

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Sprague-Dawley male rats, served as source of the S-9 fraction. A rat liver S-9 fraction buffered with 0.15M KC1. The positive control substances DMBA or 3-MCA were tested with each new batch of S-9 fraction for their ability to induce forward mutations in the CHO/HGPRT assay. Prior to use in the HGPRT test, the S-9 fraction was tested for contamination and for cytotoxicity.

Test concentrations with justification for top dose:

0.16 - 40 µg/mL (-S9)
0.11 - 60 µg/mL (+S9)
After determination of the cytotoxicity of LH 30/Z, the concentration range of LH 30/Z for the mutagenicity study was chosen ranging from approximately 0% to 90% reduction in colony forming ability.
Due to the low cytotoxicity of LH 30/Z in the mutation assays, the test article concentration was increased during the study up to 40 µg/mL without metabolic activation and up to 60.0 µg/mL with metabolic activation.

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

3-methylcholanthrene

ethylmethanesulphonate

Evaluation criteria:

An assay normally is considered acceptable for evaluation of the results only if the following
criteria are satisfied. However, the conclusion of the study will be based upon Study Director's evaluation and interpretation of the data. The activation and nonactivation assays were repeated independently in a second assay. The average cloning efficiency of the negative controls should be; at least 50%. Assays below 50% cloning efficiency will be unacceptable. The background mutant frequency (average of the negative control) should not exceed 25x10 cells. Assays with higher spontaneous mutant frequencies, however, are riot necessarily invalid if all other criteria are fulfilled.
An experimental mutant frequency is considered acceptable only if the absolute cloning efficiency is 10% or greater. The mutant frequencies for at least five treated cultures are normally determined in each assay. Mutant frequencies are normally derived from sets of 8-10 dishes for each dose level. To allow for contamination losses , an acceptable mutant frequency can be calculated from a minimum of 5 dishes. The positive control must induce a mutant frequency of at least three times that of the negative control. An assay will be considered positive if a dose-dependent and reproducible increase in mutant frequency is observed. It is desirable to obtain this dose-relation for at least 3 doses.

Statistics:

All data are presented in tabular form, descriptive statistical methods were used to calculate means and standard deviation.

Results and discussion

Test results

Key result

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

Increases in mutation frequencies were observed. However, these increases were not dose-related

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Due to the low cytotoxicity of LH 30/Z in the mutation assays, the test article concentration was increased during the study up to 40 µg/mL without metabolic activation and up to 60.0 µg/mL with metabolic activation.

Vehicle controls validity:

not specified

Untreated negative controls validity:

not examined

Remarks:

A stability test in the solvent was not done due to the non-homologous test compound.

True negative controls validity:

not specified

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

The test material, propineb was assayed for mutagenic activity at the HGPRT locus in CHO cells from 0.16 µg/ml to 40 µg/ml without activation and from 0.11 µg/ml to 60.0 µg/ml with activation.
Under both treatment conditions, cytotoxicities were induced. The absolute cloning efficiencies for the vehicle controls varied from 80.0% to 108.8% without activation and from 73.2% to 108.8% with activation demonstrating good cloning conditions for the assays.
The vehicle control mutant frequencies were all in the normal range of background frequencies for the assay. In contrast, the positive controls EMS, DMBA and 3-MCA induced a distinct mutagenic effect in mutant frequency, which was significantly increased over the
negative controls.
From the lack of dose-related and reproducible increases in mutant frequency the test material is considered nonmutagenic in the CHO-HGPRT Forward Mutation Assay, both with and without metabolic activation, according to our evaluation criteria.

Executive summary:

Propineb was evaluated for mutagenic effects at the HGPRT locus (forward mutation assay) in CHO cell cultures after in vitro treatment at concentrations up to 40.0 µg/ml (without S-9 mix) and 60.0 µg/ml (with S-9 mix). Under both treatment conditions, LH 30/Z induced cytotoxic effects as seen by decreases in relative population growth and cloning efficiency. These results revealed a significant cytotoxicity of LH 30/Z, both with and without S-9 mix.

 

There were neither dose-related nor reproducible increases in mutant frequency which were significantly elevated over the negative controls. In contrast, the positive controls ethylmethanesulfonate (without S-9 mix), 3-inethylchola-nthrene and dimethylbenzanthracene (with S-9 mix) revealed a clear mutagenic effect in the assay. From these results, the test substance LH 30/Z can be considered, as nonmutagenic in the CHO-HGPRT Forward Mutation Assay, both with and without metabolic activation.