L-Glutamine, L-alanyl-

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2008

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

475-290-9

EC Name:

-

Cas Number:

39537-23-0

Molecular formula:

C8H15N3O4

IUPAC Name:

(2R)-2-[(2S)-2-aminopropanamido]-4-carbamoylbutanoic acid

Method

Target gene:

Not applicable

Species / strain

Species / strain / cell type:

Chinese hamster lung (CHL/IU)

Details on mammalian cell type (if applicable):

For cell lines:
- Cell cycle length: 17 h

MEDIA USED
- Type and composition of media, CO2 concentration, humidity level, temperature: cells were grown in Eagle’s Minimal Essential Media with HEPES buffer and Earle’s Salts, supplemented with 10% foetal bovine serum and antibiotics and incubated at 37 °C with 5% CO2 in air

Metabolic activation:

with and without

Metabolic activation system:

Type and composition of metabolic activation system:
- source of S9: male Sprague–Dawley rats (approx. 250 g) induced with 3 consecutive daily doses of 80 mg phenobarbitone/kg bw and 100 mg beta-naphthoflavone/kg bw
- quality controls of S9: metabolic capability tested

Test concentrations with justification for top dose:

Preliminary toxicity test: 34 to 2180 µg/mL
First and second experiment: 0 to 2180 µg/mL

Vehicle / solvent:

- Vehicle/solvent used: sterile distilled water

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

mitomycin C

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION first experiment
- Exposure duration: 6 h with and without S9-mix
- Recovery phase: 18 h
- Fixation time (start of exposure up to fixation or harvest of cells): 24 h

DURATION second experiment
- Exposure duration: 24 h or 48 h without S9-mix
- Fixation time (start of exposure up to fixation or harvest of cells): 24 h or 48 h

SPINDLE INHIBITOR (cytogenetic assays): 0.1 µg/ml Colcemid two hours prior to the end of the incubation
STAIN (for cytogenetic assays): Giemsa

NUMBER OF REPLICATIONS:
- each concentration in duplicate

NUMBER OF CELLS EVALUATED:
- the first 100 consecutive well-spread metaphases from each culture to a total of 200 cells per concentration

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index calculated from 1000 cells counted & metaphases recorded

OTHER EXAMINATIONS:
- Determination of polyploidy: yes
- Determination of endoreplication: yes

Results and discussion

Test results

Key result

Species / strain:

other: Chinese Hamster Lung (CHL) cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Data on pH: no effect on pH when dosed in cell culture
- Data on osmolality: no effect on osmolality when dosed in cell culture

RANGE-FINDING/SCREENING STUDIES:
The test item at concentrations ranging from 34 to 2180 µg/mL was incubated with cell cultures for 24 and 48 h without S9-mix, and for 6 h both with and without S9-mix. The highest concentration analysed was selected based on the recommendation limit dose.

Applicant's summary and conclusion

Conclusions:

Under the test conditions applied, the test substance does not induce chromosome aberrations in cultured mammalian somatic cells, in the presence and absence of a metabolic activation system.