Praseodymium trinitrate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In an Ames test according to OECD Guideline 471, EU Method B.13/14 and EPA OPPTS 870.5100 (Thompson, 2013), praseodymium trinitrate proved to be negative for mutagenicity with and without metabolic activation.

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:

2012-10-03 - 2012-11-26

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Deviations:

no

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Version / remarks:

METI, MHLW and MAFF

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Target gene:

Salmonella typhimurium
TA1537: his C 3076; rfa-; uvrB- (frame shift mutations)
TA98: his D 3052; rfa-; uvrB-; R-factor (frame shift mutations)
TA1535: his G 46; rfa-; uvrB- (base-pair substitutions)
TA100: his G 46; rfa-; uvrB-; R-factor (base-pair substitutions)

Escherichia coli
WP2 uvrA: trp-; uvrA- (base-pair substitution)

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

S9 microsomal fraction from rats induced with phenobarbitone/beta-naphtoflavone

Test concentrations with justification for top dose:

Preliminary toxicity test: 0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 μg/plate (praseodymium trinitrate as active
ingredient).
Mutation test - experiment 1:
15, 50, 150, 500, 1500 and 5000 μg/plate (praseodymium trinitrate as active ingredient) were assayed in triplicate.

Mutation test - experiment 2: Modified following results from experiment 1:
5, 15, 50, 150, 500, 1500 and 5000 μg/plate (praseodymium trinitrate as active ingredient)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: sterile distilled water

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

2 μg/plate for WP2uvrA, 3 μg/plate for TA100, 5 μg/plate for TA1535; without S9-mix

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

80 μg/plate for TA1537; without S9 mix

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 4-Nitroquinoline-1-oxide

Remarks:

0.2 μg/plate for TA98; without S9 mix

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: other: 2-aminoanthracene

Remarks:

1 μg/plate for TA100; 2 μg/plate for TA1535 and TA1537; 10 μg/plate for WP2uvrA; with S9-mix

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Remarks:

5 μg/plate for TA98; with S9-mix

Details on test system and experimental conditions:

METHOD OF APPLICATION: plate incorporation (mutation test - experiment I) and preincubation method (mutation test - experiment II).
DURATION
- Preincubation period: 20 minutes (mutation test - experiment 2 - pre-incubation method)
- Exposure duration: 48 hours
- Selection time (if incubation with a selection agent): simultaneous with exposure
SELECTION AGENT (mutation assays): trace histidine or tryptophan supplemented
NUMBER OF REPLICATIONS: triplicate
DETERMINATION OF CYTOTOXICITY
- Method: reduction in number of revertant colonies and reduction of bacterial background lawn
- Preliminary toxicity test:
In order to select appropriate dose levels for use in the main test, a preliminary test was carried out to determine the toxicity of the test item. The concentrations tested were 0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 μg/plate (praseodymium trinitrate as active ingredient). The test was performed by mixing 0.1 mL of bacterial culture (TA100 or WP2uvrA), 2 mL of molten, trace histidine or tryptophan supplemented, top agar, 0.1 mL of test item formulation and 0.5 mL of S9-mix or phosphate buffer and overlaying onto sterile plates of Vogel-Bonner Minimal agar (30 mL/plate). Ten concentrations of the test item formulation and a vehicle control (sterile distilled water) were tested. In addition, 0.1 mL of the maximum concentration of the test item and 2 mL of molten, trace histidine or tryptophan supplemented, top agar were overlaid onto a sterile nutrient agar plate in order to assess the sterility of the test item. After approximately 48 hours incubation at 37°C the plates were assessed for numbers of revertant colonies using an automated colony counter and examined for effects on the growth of the bacterial background lawn.

Evaluation criteria:

There are several criteria for determining a positive result. Any, one, or all of the following can be used to determine the overall result of the study:
1. A dose-related increase in mutant frequency over the dose range tested
2. A reproducible increase at one or more concentrations
3. Biological relevance against in-house historical control ranges
4. Statistical analysis of data as determined by UKEMS (Mahon et al (1989))
5. Fold increase greater than two times the concurrent solvent control for any tester strain (especially if accompanied by an out-of-historical range response).
A test item will be considered non-mutagenic (negative) in the test system if the above criteria are not met. Although most experiments will give clear positive or negative results, in some instances the data generated will prohibit making a definite judgment about test item activity. Results of this type will be reported as equivocal.

Key result

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(A visible reduction in the growth of the bacterial background lawns for all of the tester strains with and without S9-mix from 1500 μg/plate was observed in both experiments)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(A visible reduction in the growth of the bacterial background lawns for all of WP2 uvr A with and without S9-mix at 5000 μg/plate was observed in experiment 1)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

The test formulation and S9-mix used in this experiment were both shown to be sterile.
Mutation test:
The test item caused a visible reduction in the growth of the bacterial background lawn and/or substantial reductions in revertant colony frequency for all of the Salmonella tester strains in the presence and absence of S9-mix at 1500 μg/plate in each experiment, and also for E. coli at the 5000 ug/plate in the presence and absence of S9-mix in the first experiment. These results were not indicative of toxicity sufficiently severe enough to prevent the test item being tested up tothe maximum recommended dose level of 5000 μg/plate. A test item precipitate (greasy and particulate in appearance) was noted at and above 1500 μg/plate in each experiment in the absence and presence of S9-mix, this observation did not prevent the scoring of revertant colonies.
No significant increases in the frequency of revertant colonies were recorded for any of the strains of bacteria, at any dose level either with or without metabolic activation or exposure method.
All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies thus confirming the activity of the S9-mix and the sensitivity of the bacterial strains.

Remarks on result:

other: all strains/cell types tested

Conclusions:

The test item, praseodymium trinitrate, was considered to be non-mutagenic under the conditions of this test.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Thompson (2013) performed an Ames (preliminary toxicity test, plate incorporation assay and preincubation method) test with S typhimurium strains TA1535, TA1537, TA98 and TA100 and Escherichia coli strain WP2uvrA in the presence and absence metabolic activation.

Following test concentrations were applied in triplicate:

Preliminary toxicity test: 0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 μg/plate (praseodymium trinitrate as active

ingredient).

Mutation test - experiment 1:

15, 50, 150, 500, 1500 and 5000 μg/plate (praseodymium trinitrate as active ingredient)

Mutation test - experiment 2: Modified following results from experiment 1:

5, 15, 50, 150, 500, 1500 and 5000 μg/plate (praseodymium trinitrate as active ingredient)

Solvent (vehicle) controls and positive controls were run in triplicate. The test item caused a visible reduction in the growth of the bacterial background lawn and/or substantial reductions in revertant colony frequency for all of the Salmonella tester strains in the presence and absence of S9-mix at 1500 μg/plate in each experiment, and also for E.coli at the 5000 ug/plate in the presence and absence of S9-mix in the first experiment. These results were not indicative of toxicity sufficiently severe enough to prevent the test item being tested up to the maximum recommended dose level of 5000 μg/plate. A test item precipitate (greasy and particulate in appearance) was noted at and above 1500 μg/plate in each experiment in the absence and presence of S9-mix, this observation did not prevent the scoring of revertant colonies.

No significant increases in the frequency of revertant colonies were recorded for any of the strains of bacteria, at any dose level either with or without metabolic activation or exposure method.

All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies thus confirming the activity of the S9-mix and the sensitivity of the bacterial strains.

Justification for classification or non-classification

Praseodymium trinitrate is not mutagenic in an Ames test. These data are conclusive but not sufficient for classification.