Trisodium dihydrogen -N,N-[bis[2-[bis(carboxylatomethyl)amino]ethyl]]glycinate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

A bacterial reverse mutation study, conducted according to OECD 471 and 472, did not cause any biologically significant increases in reverse mutation colonies and an OECD 473 in vitro chromosome abberation study in mammalian cells in a close analgue showed no evidence of clastogenic activity..

By sequestration of zinc ions, DTPA trisodium salt is expected to exert a major effect on DNA synthesis during the IGF-I stimulatory phase of the cell cycle (RS MacDonald et al 1996). The EU risk assessment of the similar chelating agent EDTA (Vol 49, 2004) concluded that on the basis of the various negative findings and the assumption of a threshold mode of action for aneugens, EDTA and its sodium salts are not mutagenic for humans. As such it is expected that DTPA and its sodium salts would not be mutagenic, based on the structural similarity of these substances. 

It is therefore concluded that DTPA trisodium salt is not mutagenic.

 

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

11 June 2001 to 19 July 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

and OECD 472

Deviations:

not specified

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

Lot No.1Y31L, 1% aqueous purity.

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:

No information

Test concentrations with justification for top dose:

Preliminary study
48.8, 195, 781, 3125, 12500 and 50000 μg/mL without S9 mix
48.8, 195, 781, 3125, 12500 and 50000 μg/mL with S9 mix
Main study
1563, 3125, 6250, 12500, 25000 and 50000 μg/mL without S9 mix
1563, 3125, 6250, 12500, 25000 and 50000 μg/mL with S9 mix

Vehicle / solvent:

Purified water

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

sodium azide

N-ethyl-N-nitro-N-nitrosoguanidine

other: 2_Aminoanthracine

Details on test system and experimental conditions:

The study was performed by the preincubation method. Two plates were used for each test, and each dose level.
1)Dose setting study and main study
In the dose setting study, bacterial suspensions of respective strains dispensed and stored frozen were thawed and 20 μL each was inoculated in 10 mL of nutrient broth culture solution and incubated by shaking (65 reciprocations/minute) at 37°C in a dark room for 12 hours. A 0.1-mL aliquot of this culture solution, 0.5 mL of S9 mix (0.5 mL of 0.1M sodium phosphate buffer, pH 7.4, when S9 mix was not used) and 0.1 mL of 48.8, 195, 781, 3125, 12500 or 50000 μg/mL test substance solution (4.88, 19.5, 78.1, 312.5, 1250 or 5000 μg/plate) were measured in a sterilized test tube and preincubated at 37°C for 20 minutes. Then, 2 mL of the overlay agar containing amino acid was added, mixed to spread over the minimum glucose agar plate medium, and incubated at 37°C for 48 hours. After incubation, reverse mutant colonies were counted macroscopically and bacterial growth inhibition was observed under a stereomicroscope.
The main study was conducted in the same manner as in the dose setting study. Based on the results of the dose setting study, 6 concentrations (156.3, 312.5, 625, 1250, 2500 and 5000 μg/plate) were provided with 50000 μg/mL (= 5000 μg/plate) as the highest concentration and common ratio of 2.
A vehicle control and a positive control were provided for each bacterial strain and tested in the same manner.
2)Sterility tests for S9 mix and the test substance solution
On a plate of normal agar plate medium (20 mL) prepared using the normal agar plate medium (Eiken Chemical Co., Ltd., pH 7.2), 0.1 mL each of S9 mix, 0.1M sodium phosphate buffer, 2 types of overlay agar and the test substance solution of each concentration were spread and incubated at 37°C for 48 hours.

Evaluation criteria:

When the mean value of 2 plates obtained with the test substance is compared to the data obtained with the vehicle control and the positive control, the result is judged as positive if the number of reverse mutant colonies is twice or more that of the vehicle control and the result is dose-dependence and reproducible.

Statistics:

Not specified

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:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

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:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

The number of reverse mutant colonies in the positive control substance was twice or more that in the vehicle control for all strains both without S9 mix system and with S9 mix in the dose setting study and the main study, demonstrating mutagenicity.

In the sterility test, microbial contamination was not detected in any of the test solutions.

The dose setting study was conducted at six dose levels with 5000 μg/plate at the highest dose level and common ratio of four for both without S9 mix and with S9 mix.  Growth inhibition was observed at 5000 μg/plate for all bacterial strains except for TA98 in wihtout S9 mix and for TA100, WP2uvrA and TA1537 with S9 mix. The number of reverse mutant colonies was not twice or more than that of the vehicle control.

The main study was conducted at six dose levels with 5000 μg/plate at the highest dose level and common ratio of two for both without S9 mix and with S9 mix, based on the results of the dose setting study. Growth inhibition was observed at 5000 μg/plate for TA100 and TA1537 without S9 mix and for TA100, WP2uvrA and TA1537 with S9 mix. The number of reverse mutant colonies was not twice or more than that of the vehicle control for all bacterial strains

Conclusions:

The bacterial reverse mutation study of DTPA did not show the number of reverse mutant colonies twice or more that in the vehicle control even at the highest dose level of 5000 μg/plate in the dose setting study and the main study with or without metabolic activation. Growth inhibition was noted in some of the bacterial strains.
Based on the above results, it was concluded that the test substance was not mutagenic.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Justification for classification or non-classification

DTPA trisodium salt is considered not to meet the CLP criteria for classification for mutagenicity.