Disodium L-cystinate

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:

1983

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

publication

Title:

Mutagenicity of Glutathione and Cysteine in the Ames Test

Author:

Glatt H, Protic-Sabljic M, Oesch F

Year:

1983

Bibliographic source:

Science 220 (4600), 961-963

Report date:

1983

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

- Principle of test: Ames test (Salmonella typhimurium reverse mutation assay) with histidine-dependent strain.
- Short description of test conditions: Postmitochondrial supernatant was prepared from homogenates of kidney and lvier from untreated adult male Sprague-Dawley rats. Histidine-dependent bacteria of Salmonella typhimurium TA100, the subcellular fraction from 100 mg of tissue with or without an NADPH-generating system, a neutralized solution of the test compound in water, and histidine-poor soft agar were mixed and added to culture plates containing minimal agar. The colonies that reverted to histidine independence were counted after 2 days' incubation in the dark.
- Parameters analysed / observed: Number of revertants per plate

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Cystine

EC Number:

200-296-3

EC Name:

Cystine

Cas Number:

56-89-3

Molecular formula:

C6H12N2O4S2

IUPAC Name:

cystine

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

Cysteine sourced from three different manufacturers: Boehringer, Sigma and Merck

Method

Target gene:

Histidine

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:

S. typhimurium TA 1538

Metabolic activation:

with and without

Metabolic activation system:

liver or kidney S9 (postmitochondrial supernatant)

Test concentrations with justification for top dose:

0, 5, 10, 20, 40 mM (according to figure)

Vehicle / solvent:

not specified

Controls

Untreated negative controls:

yes

Remarks:

Hydrogen peroxide

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar

DURATION
- Preincubation period: not specified
- Exposure duration: 2 days
- Expression time (cells in growth medium): not specified
- Selection time (if incubation with a selection agent): 2 days

NUMBER OF REPLICATIONS: 3

Rationale for test conditions:

Followed the Ames test procedure.

Evaluation criteria:

Counted the number of histidine revertants.

Statistics:

Not specified

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

valid

Remarks:

Hydrogen peroxide

Positive controls validity:

not specified

Test results 2

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

valid

Remarks:

hydrogen peroxide

Positive controls validity:

not specified

Test results 3

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

valid

Remarks:

hydrogen peroxide

Positive controls validity:

not specified

Test results 4

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

valid

Remarks:

hydrogen perioxide

Positive controls validity:

not specified

Test results 5

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

valid

Remarks:

hydrogen perioxide

Positive controls validity:

not specified

Test results 6

Key result

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

valid

Remarks:

hydrogen perioxide

Positive controls validity:

not specified

Additional information on results:

S. typhimurium TA 98, TA 1535, TA 1539, TA 1538: negative, with and without metabolic activation

S. typhimurium TA100:
In the absence of S9, cysteine did not increase the number of revertants.
With the addition of liver S9, cysteine increased the number of revertants
With the addition of kidney S9, cysteine increased the number of revertants severalfold above the spontaneous level.
Different batches from different manufacturers (Boehringer, Sigma, Merk) did not show any quantitative differences.

Applicant's summary and conclusion

Conclusions:

In the presence of liver or kidney S9, cysteine increased the number of histidine revertants in S typhimurium TA 100. Without S9, no increase was observed.

In S. typhimurium TA 98, TA 1535, TA 1539, TA 1538, in increase in the number of histidine revertants was reported, with or without metabolic activation.

Executive summary:

In an Ames assay testing up to 40 mM of cysteine, the postmitochondrial supernatant from rat liver and kidney homogenates induced test substance cysteine to revert Salmonella typhimurium TA100 to histidine independence. Cysteine exhibited mutagenic properties in the presence of S9 liver and kidney metabolic activation in Salmonella typhimurium TA100. In S. typhimurium  TA 98, TA 1535, TA 1539, TA 1538, in increase in the number of histidine revertants was reported, with or without metabolic activation.

The increased number of mutants in the presence of relatively high concentration of physiological compounds could be the result not of mutagenicity but of nutritional interactions between these compounds and histidine because the histidine-dependent strains TA1537, TA1538, TA98 and TA1535 which differ from TA100 in the mutation leading to histidine dependence or in the DNA repair system, showed either no revertants or very low numbers of revertants in response to glutathione and cysteine compared with the number induced in strain TA100.