Vinyl ethylene carbonate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 03 Sep to 12 Oct 2021

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2022

Report date:

2022

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosomal Aberration Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

(4R)-4-ethenyl-1,3-dioxolan-2-one; (4S)-4-ethenyl-1,3-dioxolan-2-one

EC Number:

700-261-7

Cas Number:

4427-96-7

Molecular formula:

C5H6O3

IUPAC Name:

(4R)-4-ethenyl-1,3-dioxolan-2-one; (4S)-4-ethenyl-1,3-dioxolan-2-one

Test material form:

liquid

Specific details on test material used for the study:

Batch No.: 10121040902
Purity: 99.92%

Method

Species / strain

Species / strain / cell type:

Chinese hamster lung (CHL/IU)

Metabolic activation:

with and without

Metabolic activation system:

Type and composition of metabolic activation system:
- source of S9: Aroclor 1254 induced rat-liver S9
- method of preparation of S9 mix: The S9 was prepared from the livers of the rats induced with Aroclor1254, and the details were shown below: Male SD (Sprague-Dawley) rats were treated by a single intraperitoneal injection of Aroclor 1254 at the dose of 500 mg/kg body weight five days prior to the S9 preparation. The livers of the rats were taken out under aseptic conditions, homogenized in a 0.15 mol/L KCl solution (1g liver: 3ml KCl solution) and centrifuged at 11000rpm for 10 min with AllegraTM 64R Auto Freeze Centrifuge. The supernatant of S9 was stored in liquid nitrogen at approximate -196℃ for use not more than one year.

Test concentrations with justification for top dose:

6 doses of 2000, 800, 320, 128, 51.2 and 20.48 μg/ml

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO

- Justification for choice of solvent/vehicle: The test results of test item solubility showed that the test item could be dissolved in DMSO at the concentration of 400mg/ml, so DMSO was used as solvent.

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

ethylmethanesulphonate

Remarks:

Used as the positive control in the absence of S9 mix (S9-)

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: Cyclophosphamide monohydrate

Remarks:

Used as the positive control in the presence of S9 mix (S9+)

Details on test system and experimental conditions:

NUMBER OF REPLICATIONS:
- Number of cultures per concentration (single, duplicate, triplicate): duplicate
- Number of independent experiments: 1

METHOD OF TREATMENT/ EXPOSURE:
- Cell density at seeding (if applicable): 2.0E+05 CHL cells
- Test substance added in medium

TREATMENT AND HARVEST SCHEDULE:
- Preincubation period, if applicable:
- Exposure duration/duration of treatment: 3h or 24h
- Harvest time after the end of treatment (sampling/recovery times): 23h

FOR CHROMOSOME ABERRATION:
Based on the results of cytotoxicity in each treatment condition, the cells of the doses of 2000, 800 and 320μg/ml exposed in the above three treatment conditions and all controls were chosen for chromosome preparation.
300 well-spread metaphase cells equally divided among the duplicates were scored per dose and control for each treatment condition. Different types of structural chromosome aberrations listed below were scored. Gaps were scored separately. Polyploidy and endoreduplication were scored at the same time. The number of centromeres equal to the modal number ± 2 should be met.

Evaluation criteria:

Providing that all validity criteria are fulfilled in any of the treatment condition examined, when all of the following criteria are met, the result is considered to be clearly positive:
a) At least one of the test dose exhibits a statistically significant increase compared with the concurrent solvent control (P＜0.05);
b) The increase is dose-related when evaluated with an appropriate trend test.
c) Any of the results is outside the 95% control limits of historical negative control data distribution in this lab.

Providing that all validity criteria are fulfilled in all of the treatment conditions examined, when all of the following criteria are met, the result is considered to be clearly negative:
a) None of the test dose exhibits a statistically significant increase compared with the concurrent negative control;
b) There is no dose-related increase when evaluated with an appropriate trend test. c) All results of the doses are inside the 95% control limits of historical negative control data distribution in this lab.

Results and discussion

Test results

Key result

Species / strain:

Chinese hamster lung (CHL/IU)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

The results of test item precipitate: In this test, no test item precipitate was observed at all designed dose levels at the beginning and end of the treatment in three treatment conditions.

The results for cytotoxicity: The RICC of the cells exposed for 3h in the absence of S9 mix at the doses of 2000, 800, 320, 128, 51.2 and 20.48μg/ml were 78%, 84%, 92%, 94%, 98% and 102%; the RICC of the cells exposed for 3h in the presence of S9 mix at the doses of 2000, 800, 320, 128, 51.2 and 20.48μg/ml were 83%, 95%, 91%, 94%, 102% and 100%; the RICC of the cells exposed for 24h in the absence of S9 mix at the doses of 2000, 800, 320, 128, 51.2 and 20.48μg/ml were 81%, 92%, 97%, 94%, 91% and 95%.

The results of the microscopic analysis: the percentage of cells with structural chromosomal aberration excluding gap exposed for 3h in the absence of S9 mix at the doses of 2000, 800 and 320μg/ml were 0.7%, 0.3% and 0.3%; the percentage of cells with structural chromosomal aberration excluding gap exposed for 3h in the presence of S9 mix at the doses of 2000, 800 and 320μg/ml were 0.7%, 0.3% and 0.3%; the percentage of cells with structural chromosomal aberration excluding gap exposed for 24h in the absence of S9 mix at the doses of 2000, 800 and 320μg/ml were 0.7%, 0.7% and 0.3%. No statistically significant difference was observed at all doses of each test condition as compared with the concurrent solvent control (P>0.05). At the same time, statistically significant increases were observed in the cells of all positive controls as compared with the concurrent solvent controls(P<0.01).

Applicant's summary and conclusion

Conclusions:

The results of chromosomal aberration test were negative in three treatment conditions. Thus the test item was considered not to cause structural chromosomal aberrations in cultured mammalian cells.

Executive summary:

The study was performed to evaluate test item for its capability to cause structural chromosomal aberrations in cultured mammalian cells. Three treatment conditions were performed in this study including exposing to the test item for 3 hours with and without the metabolic activation, and for 24 hours without the metabolic activation. The method of this study was designed to be in compliance with the OECD Guideline 473.

In this study, the results of chromosomal aberration test were negative in three treatment conditions. Thus the test item, was considered not to cause structural chromosomal aberrations in cultured mammalian cells under the conditions of this study.