Sodium O-isobutyl dithiocarbonate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

31.05.2021-15.07.2021

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Performed to guidelines

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2021

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

the study was carried out in accordance with microplate format of the Ames fluctuation assay, which differs from the methods described in details in the OECD 471 Guideline and EU Method B.13/14.

GLP compliance:

yes (incl. QA statement)

Remarks:

22/2020/DPL

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Sodium O-isobutyl dithiocarbonate

EC Number:

246-805-2

EC Name:

Sodium O-isobutyl dithiocarbonate

Cas Number:

25306-75-6

Molecular formula:

C5H10OS2.Na

IUPAC Name:

sodium [(2-methylpropoxy)methanethioyl]sulfanide

Constituent 2

Reference substance name:

Sodium isobutyl xanthate

IUPAC Name:

Sodium isobutyl xanthate

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

94±1%

Method

Species / strain

Species / strain 1

Species / strain / cell type:

E. coli WP2 uvr A pKM 101

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

The following components from Xenometrix were used to prepare the metabolic activity system:
◆ phenobarbital/β-naphtoflavone induced S9 fraction (cat. no. PRS-PB02, lot. no. F1903114). The Certificate of Analysis of Phenobarbital/β-naphtoflavone induced S9 – Postmitochondiral Supernatant was presented in Appendix no. 5.
◆ S9-Buffer Salts (cat. no. PCO-0800, lot. no. ND09648P), containing 0.20 M NaH2PO4, and 0.25 M MgCl*6H2O and 1 M KCl.
◆ 0.04 M S9-NADP (cat. no. PCO-0830, lot. no. X1904SNAA),
The metabolic activity system was supplemented with 0.20 M glucose-6-phosphatase solution (G-6-P; Sigma-Aldrich, cat. no. G7879, lot. no. SLBZ7814).
The defrosted reagents were stored on ice. To prepare 30 % mixture of the S9 fraction, the following volumes of the reagents were mixed (for 5 strains): 3.106 mL S9 – buffer – salts, 0.136 mL G-6-P, 0.540 mL S9 – NADP and 1.620 mL phenobarbital/β-naphtoflavone induced S9 fraction. This mixture was prepared immediately before use and added to the bacteria in the exposure media. The final concentration of S9 in culture was 4.5%.
The S9 fraction can be toxic for Escherichia coli strains. So, the number of positive wells may be lower than in other strains

Test concentrations with justification for top dose:

5.000 mg/mL, 1.581 mg/mL, 0.500 mg/mL, 0.158 mg/mL, 0.050 mg/mL, 0.016 mg/mL.
There were three replicates of each concentration.

Vehicle / solvent:

Water

Controls

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

2-nitrofluorene

other: N4-aminocitidine, 2-aminoanthracene

Results and discussion

Test results

Test results 1

Species / strain:

E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

True negative controls validity:

not applicable

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:

no cytotoxicity

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

True negative controls validity:

not applicable

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

True negative controls validity:

not applicable

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

True negative controls validity:

not applicable

Positive controls validity:

valid

Test results 5

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

True negative controls validity:

not applicable

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

The test item (Sodium isobutyl xanthate (Sodium O-isobutyl dithiocarbonate)) was tested in Bacterial Reverse Mutation Test to indicate potential mutagenic risk. The bacterial reverse mutation test is commonly employed as an initial screen for genotoxic activity and for point mutation-inducing activity. There was demonstrated that many chemicals that are positive in this test also exhibit mutagenic activity in other tests, but the correlation is not absolute [5]. The test was performed with Ames MPFTM Penta I Microplate Format Mutagenicity Assay kit (Xenometrix).
Statistically significant difference (p<0.05) was noticed in only one result. This result was also outside the historical data of Negative Control. However, the average number of positive wells were not bigger than twice the baseline value and no concentration-response correlation was observed (r≥0.75). Therefore, genotoxic properties of the test item were not detected.
Hence, it may be concluded that the test item under the test conditions is not a mutagenic in the tested species.