Zirconium dinitrate oxide

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015.01.09 - 2015.04.19

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

As the study was designed as a combined range finder/limit test, the testing laboratory performed it under non-GLP conditions as it is currently done for range-finding studies. However despite the fact that the study is not GLP compliant, it is well performed and the results are considered valid and of good quality.

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

GLP compliance:

no

Remarks:

The biological study was not GLP-compliant. Measurements of samples from the prel. experiments were analysed outside GLP. By contrast, the measurements of samples from the main study were conducted under GLP conditions including method validation.

Analytical monitoring:

yes

Details on sampling:

Stirring experiments:
Due to the low solubility of the test item in test medium, stirring experiments were performed in order to: (1) check whether it was possible to obtain zirconium levels in solution that are well above the limit of quantification (LOQ) of the analytical method, and (2) to determine the stirring time allowing an optimal dissolution of zirconium during the test. Sampling conditions were as follows during the stirring experiments:

- At time 0 and after 24 and 96 h of stirring, samples were taken from the test solution with nominal loading rate 100 mg/L (anhydrous test substance). In case pH adjustment was performed, samples were taken after pH adjustment.
- All samples were taken in plastic tubes and were membrane-filtered (0.22 µm Millipore Express PLUS(PES) membrane filter).
- Samples were acidified with HNO3 (2% v/v) and stored in the refrigerator before analysis. Samples to be analyzed were sent at ambient temperature (2-8°C) to the analytical laboratory (Seibersdorf Labor GmbH, Austria), where they arrived in good condition.

Combined range finder/limit test:
- Samples were taken from the control and from the 1, 10 and 100% saturated test solutions at the start and end of each renewal period (medium renewal after 48 h), both from freshly prepared and aged test media.
- The sampling procedure and treatment was the same as described above for the stirring experiments.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
First stirring experiment:
- A stock solution with a nominal loading rate of 100 mg/L (anhydrous test substance) was prepared by adding 73.6 mg test item in 500 mL aquarium water. The test item was mixed into the test water as homogeneously as possible using intense stirring. After initial stirring, the pH was measured as 7.23. After 3 hours the pH was checked again, the measured value was 7.54. After 24 hours, a pH of 8.10 was measured and while stirring adjusted (using HCl) to the value measured at the start (7.23). At the end of the experiment (96 hours) the pH was measured again (7.89) and adjusted to 7.23.

Second stirring experiment:
- A second stirring experiment was performed with adjustment of pH to 6.0. This was done because greater quantities of dissolved zirconium are expected at this pH, whilst still remaining within the physiologically acceptable range for rainbow trout. Preparation of the stock solution with a nominal loading rate of 100 mg/L (anhydrous test substance) was the same as described for the first stirring experiment. After initial stirring, the pH was measured (6.62) and then adjusted (using HCl) to 6.00. After 24 hours the pH was checked again and determined to be 7.35, then it was adjusted to 6.00 again while stirring. The pH was measured after 96 hours (7.02) and then it was adjusted again to 6.00.

Combined range finder/limit test:
- A stock solution with nominal loading rate 100 mg/L (anhydrous zirconium dinitrate oxide) was prepared by adding 2 x 625 mg test substance to 2 x 4250 mL test medium followed by 24 h stirring. The second stirring experiment had shown that the highest zirconium concentration in solution was obtained after 24 h stirring. After 24 h stirring, pH was measured (6.64) and adjusted (using HCl) to 6.00. Then the stock solution was filtered and the filtrate (i.e. the 100% saturated solution) was used as highest concentrated test medium. The filtrate was diluted with test water to obtain the test media of the additional lower saturated solutions (1% and 10%, v/v).

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: Rainbow trout (Oncorhynchus mykiss)
- Source: HOITSY & RIEGER Kft., Trout Farm, Miskolc-Lillafüred, Hungary
- Length at study initiation (length definition, mean, range and SD): 4.0 – 4.2 cm
- Body weight at study initiation: 0.66-0.96 g (calculated mean weight of 1 fish)
- Method of breeding: Fish were bred in a well-known fish farm, under disease- and parasite-controlled conditions.
- Feeding during test: no

ACCLIMATION
- Acclimation period: more than 12 days (~ 3 months)
- Acclimation conditions (same as test or not): same as test
- Type and amount of food: not reported
- Feeding frequency: not reported
- Health during acclimation (any mortality observed): Fish were observed during the acclimatisation and test periods, the fish were healthy. Mortality was less than 5% of the population for seven days before the start the test.

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

241 mg/L CaCO3

Test temperature:

15.7 °C - 16.8 °C

pH:

0 h (fresh media): 6.18-6.59
24 h (aged media): 7.24-7.34
48 h (aged media): 7.41-7.85
48 h (fresh media): 6.10-6.62
72 h (aged media): 7.43-7.92
96 h (aged media): 7.10-7.36

Dissolved oxygen:

not measured

Salinity:

not applicable

Nominal and measured concentrations:

* 1st stirring experiment (nominal loading rate 100 mg/L (anhydrous test compound)):
Measured concentration at time = 0h:
control: < 0.03 mg Zr/L; loading rate of 100 mg Zr/L: 0.03 mg Zr/L
Measured concentration at time = 24h:
loading rate of 100 mg Zr/L: < 0.03 mg Zr/L before and after pH adjustment
Measured concentration at time = 96 h:
loading rate of 100 mg Zr/L: 24 mg Zr/L before pH adjustment; after pH adjustment: 0.62 mg Zr/L

* 2nd stirring experiment (same loading rate):
Measured concentration at time = 0h:
control: < 0.03 mg Zr/L; loading rate of 100 mg Zr/L: 1.7 mg Zr/L before pH adjustment; after pH adjustment: 0.03 mg Zr/L
Measured concentration at time = 24h:
loading rate of 100 mg Zr/L: 0.26 mg Zr/L before pH adjustment; after pH adjustment: 1.10 mg Zr/L
Measured concentration at time = 96 h:
loading rate of 100 mg Zr/L: 0.49 mg Zr/L before pH adjustment; after pH adjustment: 0.79 mg Zr/L

* main test (same loading rate corresponding to a 100% saturated solution and 1% and 10% dilutions v/v)
All concentrations were below 0.03 mg Zr/L, no dissolved zirconium was observed above the limit of quantification in any of the samples.

Details on test conditions:

TEST SYSTEM
- Test vessel: aquarium
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: glass, fill volume 8 L for the 100% saturated test solution and the control and 3 L for the 1% and 10% saturated solution
- Aeration: not specified
- Renewal rate of test solution (frequency/flow rate): after 48 h (semi-static test)
- No. of organisms per vessel: 7 fish for the 100% saturated test solution and the control, 3 fish for the 1% and 10% saturated test solutions
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.66-0.84 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The dilution water was circulated, cooled and filtered water. Source not specified.

OTHER TEST CONDITIONS
- Adjustment of pH: yes, to 6.0
- Photoperiod: 16 hours light and 8 hours darkness.
- Light intensity: not specified

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): mortality after 3, 6, 24, 48, 72 and 96 hours

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10
- Justification for using less concentrations than requested by guideline: combined range finder/limit test was performed because no adverse effects were expected
- Range finding study: the study was a combined range finder/limit test
- Test concentrations: 1, 10 and 100% saturated solutions

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

anhydrous form

Basis for effect:

mortality (fish)

Details on results:

No mortalities were observed in any of the test solutions during the combined range finder/limit test.
No dissolved Zr was observed > LOQ (0.030 mg Zr/L) in any of the test solutions.
In the absence of adverse effects, and because no dissolved Zr could be obtained in the test solutions (i.e., absence of relevant exposure), it was decided not to perform a definitive study. The results of the non-GLP combined range finder/limit test can be used for drawing conclusions.

Reported statistics and error estimates:

Not applicable (no mortalities or other adverse effects observed).

Validity criteria fulfilled:

yes

Conclusions:

The 96-h acute toxicity of zirconium dinitrate oxide to rainbow trout (Oncorhynchus mykiss) was studied under semi-static conditions in a combined range finder/limit test, according to OECD guideline 203. The test animals were exposed to 1, 10 and 100% saturated solutions. A nominal loading rate of 100 mg/L (anhydrous test substance) was used for preparation of the 100% saturated solution. The pH of the initial solution was adjusted to 6.0 in order to try to maximise quantities of dissolved zirconium whilst reamining a physiologically acceptable pH level for rainbow trout. The solution was then stirred for 24 h, after which pH was again adjusted, and the test solution was filtered. The filtrate was then further diluted to give 1 and 10% v/v dilutions of the saturated solution. No mortality was observed during the test, neither in the control nor in the groups exposed to the test item. The 96-h LL50, being the loading rate killing 50% of the fish, was thus set > 100 mg/L (nominal loading rate, referring to anhydrous zirconium dinitrate oxide).

Reference 2

Endpoint:

short-term toxicity to fish

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Description of key information

The endpoint is covered by a study performed with zirconium dinitrate oxide (Sipos, 2015) reporting a 96-h LL50 > 100 mg/L (anhydrous zirconium dinitrate oxide) for rainbow trout. In this study, no measurable dissolved zirconium levels could be obtained, even at a pH of 6.0, which was considered as the lowest pH feasible for testing with rainbow trout. Based on these findings, zirconium dinitrate oxide is concluded to be not harmful or toxic to fish.

Additional information

The recent study of Sipos (2015), investigating the acute toxicity of zirconium dinitrate oxide in rainbow trout (Oncorhynchus mykiss) according to OECD guideline 203, was considered as the key study for endpoint coverage. In this study, a combined range finder/limit test was performed with 1, 10 and 100% saturated solutions. The 100% saturated solution was prepared using a nominal loading rate of 100 mg/L (anhydrous zirconium dinitrate oxide). The pH of the initial solution was adjusted to 6.0 in order to try to maximise quantities of dissolved zirconium, while still remaining a physiologically acceptable pH level for rainbow trout. After intense stirring for 24 h, pH was adjusted again to 6.0 and the solution was filtered. The filtrate was then further diluted to give 1 and 10% v/v dilutions of the saturated solution. Dissolved zirconium concentrations above the LOQ of 0.03 mg Zr/L could not be detected in any of the solutions throughout the test. In addition, no mortality of fish was observed after 96 h in any of the test solutions. Based on the results of this study, the 96-h LL50 was concluded to be > 100 mg/L (anhydrous zirconium dinitrate oxide).