L-(+)-2,5-diaminopentanoic acid

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

L-(+)-2,5-diaminopentanoic acid does not exhibit genetic toxicity.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: Japanese Guidelines for Genotoxicity Studies of Drugs and the Standards for Mutagenicity Tests Using Microorganisms

Deviations:

no

GLP compliance:

yes

Type of assay:

other: in vitro mammalian chromosome aberration test

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Kyowa Hakko Bio Co., Ltd. Batches 080926 and 083019
- Expiration date of the lot/batch: practically stable
- Purity test date: 1993-08-27
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: stable
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: highly soluble
-
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: none
Purity >/= 99.9 %

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

Preliminary test: 13, 26, 53, 105, 211, 422, 843, or 1686 µg/mL
Justification: Guidelines

Short-term exposure experiments: 422, 843, or 1686 µg/mL.
Justification: 1686 µg/mLnegative in preliminary test.

Continuous experiments: 422, 843, and 1686 µg/mL
Justification: 1686 µg/mLnegative in preliminary test.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: physiological saline

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

mitomycin C

Details on test system and experimental conditions:

A preliminary cell growth inhibition test was conducted in order to select the concentrations of L-ornithine monohydrochloride to be used in the chromosome aberration test. In accordance with the Japanese Ministry of Labor guidelines, the top dose tested in the range finding assay was 0.01 M (equivalent to 1686 µg/mL). Briefly, 2 x 10exp4 cells were seeded into a 60 mm culture dish and incubated for 3 days. The test article was applied to the culture medium at concentrations of 0 (solvent control) 13, 26, 53, 105, 211, 422, 843, or 1686 µg/mL and cells were incubated for either 24 or 48 h at 37 °C. The procedure was repeated with the addition of S9 mix. Cell growth inhibition rate was measured as a percentage of cells in the solvent control.

In the short-term exposure experiments, L-ornithine was dissolved in physiological saline and added to the culture media at final concentrations of 422, 843, or 1686 µg/mL, in the presence or absence of exogenous metabolic activation (S9 mix). Untreated cultures and cultures incubated with saline were used as the negative controls, and 0.10 µg/mL mitomycin C (absence of exogenous metabolic activation) or 7.0 lg/mL benzo[a]pyrene (presence of exogenous metabolic activation) were used as positive controls. Cells were incubated with the solution containing the test article or controls for 6 h at 37 °C, followed by a rinse and further incubation in untreated culture media for an additional 18 h at 37 °C (recovery period). Colcemid (0.2 µg/mL) was added to the dishes 2 h before the end of the recovery period.

In the continuous experiments, cells were incubated with the solution containing the test article at final concentrations of 422, 843, and 1686 µg/mL for 24 or 48 h in the absence of metabolic activation. Untreated cultures and cultures incubated with saline were used as negative controls, and 0.05 µg/mL of mitomycin C was used as a positive control. Colcemid (0.2 µg/mL) was added to the dishes 2 h prior to the end of the treatment period. All cultures were prepared in duplicate.

Evaluation criteria:

The frequency of chromosome aberrations or polyploidy did not exceed 5% at any of the concentrations of L-ornithine monohydrochloride tested, in either the short-term or continuous experiments. The frequency of chromosome aberrations in cells treated with mitomycin C or benzo[a]pyrene were consistent with historical positive control data.

Key result

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with

Genotoxicity:

not determined

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Conclusions:

Under the conditions of the experiment, it was concluded that L-(+)-2,5-diaminopentanoic acid did not induce chromosome aberrations or polyploidy.

Executive summary:

L-(+)-2,5-diaminopentanoic acid was tested for genetic toxicity in an in vitro cytogenicity study in mammalian cells.

No evidence of genotoxicity was observed in the chromosome aberration test at doses of up to 1686 mg/mL, both in the presence and absence of metabolic activation.

Reference 2

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: Japanese Guidelines for Genotoxicity Studies of Drugs and the Standards for Mutagenicity Tests Using Microorganisms

Version / remarks:

Ministry of Labour, 1997; MHW, 1999

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Kyowa Hakko Bio Co., Ltd. Batches 080926 and 083019
- Expiration date of the lot/batch: practically stable
- Purity test date: 1993-08-27
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: stable
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: highly soluble
-
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: none
Purity >/= 99.9 %

Target gene:

uvrA

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:

S9 mix

Test concentrations with justification for top dose:

1.2, 4.9, 20, 78, 313, 1250, or 5000 µg/plate of L-ornithine monohydrochloride dissolved in water. No inhibition of growth was observed in any of the tester strains in the presence and absence of exogenous metabolic activation (S9 mix); therefore, a concentration of 5000 µg/plate was selected as the highest dose level tested in the main study.
Main study: 313, 625, 1250, 2500 or 5000 µg/plate in the absence and presence of exogenous metabolic activation.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle: L-Orn-HCl is well soluble in water

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

sodium azide

benzo(a)pyrene

other: 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide (AF-2), 2-methoxy-6-chloro- 9-[3-(2-chloroethyl)aminopropylamino]acridine 2HCl (ICR-191), 2-aminoanthracene.

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 20 min
- Exposure duration: 48 h
- Expression time (cells in growth medium): 48 h

NUMBER OF REPLICATIONS: 2

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED: Volony Analyzer CA-11

Evaluation criteria:

The test item was considered mutagenic if a greater than 2-fold increase in the number of revertant colonies compared to the water control occurred, and a dose response and reproducibility were observed.

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Conclusions:

Under the conditions of the experiment, it was concluded that L-(+)-2,5-diaminopentanoic acid was not mutagenic in the bacterial reverse mutation assay.

Executive summary:

L-(+)-2,5-diaminopentanoic acid was tested by the Ames-test with and without metabolic activation.

No evidence of genotoxicity was observed in the reverse bacterial mutation assay at doses of up to 5000 µg/plate, both in the presence and absence of metabolic activation.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Justification for classification or non-classification

L-(+)-2,5-diaminopentanoic acid does not exhibit genetic toxicity.