oxozirconium; silane

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

2018-01-08 to 2018-03-09

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Sampling method: Two samples were taken from the control and the 100% v/v saturated solution test group from the bulk test preparation at 0 hours and from the pooled replicates at 72 hours for immediate quantitative analysis (dissolved silicon).
- Sample storage conditions before analysis: The 0-h and 72-h samples were analysed on the day of receipt.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
*Stirring experiment:
A nominal amount of test item (500 mg) was dispersed in 5 liters of culture medium with the aid of vigorous magnetic stirring for 96 hours. After 3, 24 and 96 hours, the pH of the media was checked, adjusted to pH 7.5 +/- 0.1, and a sample was taken for chemical analysis (i.e. dissolved silicon) after filtration through a 0.2 µm Gelman Acrocap filter (initial 100 mL discarded). Dissolved silicon concentrations were 0.138, 0.366 and 0.597 mg Si/L after 3, 24 and 96 hours of stirring, respectively. Based on these results, the 96-h stirring period was selected for the definitive test.

*Definitive test:
A nominal amount of test item (500 mg) was dispersed in 5 liters of culture medium with the aid of vigorous magnetic stirring for 96 hours. After 96 hours the stirring was stopped and any undissolved test item was removed by filtration through a 0.2 μm Gelman Acrocap filter (first approximate 100 mL discarded in order to pre-condition the filter) to give a 100% v/v saturated solution. An aliquot (1 liter) of this stock solution was inoculated with 5.3 mL of algal suspension to give the required test concentration of 100% v/v saturated solution.
The stock solution and the prepared concentration were inverted several times to ensure adequate mixing and homogeneity.

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Strain: CCAP 278/4
- Source (laboratory, culture collection): Liquid cultures of Pseudokirchneriella subcapitata were obtained from the Culture Collection of Algae and Protozoa (CCAP), SAMS Research Services Ltd, Scottish Marine Institute, Oban, Argyll, Scotland.
- Method of cultivation: Master cultures were maintained in the laboratory by the periodic replenishment of culture medium. The master cultures were kept under constant agitation by orbital shaker (approximately 150 rpm) and constant illumination at 24 ± 1°C.
- Prior to the start of the test sufficient master culture was added to approximately 100 mL volumes of culture media contained in conical flasks to give an initial cell density of approximately 10^3 cells/mL. The flasks were plugged with polyurethane foam stoppers and kept under constant agitation by orbital shaker (approximately 150 rpm) and constant illumination at 24 ± 1°C until the algal cell density was approximately 10^4 to 10^5 cells/mL. These pre-cultures were used for inoculation of the test solutions at an initial cell density of 5E+03 cells/mL.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Hardness:

14.8 mg CaCO3/L

Test temperature:

24 ± °1C

pH:

t=0h: 7.7
t=72h: 7.9-8.3

Dissolved oxygen:

not applicable

Salinity:

not applicable

Nominal and measured concentrations:

Range finder:
Nominal concentrations: 0, 0.10, 1.0, 10 and 100% v/v saturated solution (with nominal loading rate of 100 mg/L)
Measured concentrations of silicon (mg/L) at t=0h: Measured concentrations of silicon (mg/L) at t=72h:
Final test:
Nominal concentrations: 0 and 100% v/v saturated solution (with nominal loading rate of 100 mg/L)
Measured concentrations of silicon (mg/L) at t=0h: Measured concentrations of silicon (mg/L) at t=72h:

Details on test conditions:

TEST SYSTEM
- Test vessel: 250-mL glass conical flasks
- Type (delete if not applicable): plugged with polyurethane foam bungs
- Material, size, headspace, fill volume: 250-mL glass conical flasks filled with 100 mL test solution
- Initial cells density: 5,000 cells/mL
- Control end cells density: 1.56 x 10^6 cells/mL
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes
* NaNO3: 25.5 mg/L
* MgCl2.6H2O: 12.16 mg/L
* CaCl2.2H2O: 4.41 mg/L
* MgSO4.7H2O: 14.6 mg/L
* K2HPO4: 1.044 mg/L
* NaHCO3: 15.0 mg/L
* H3BO3: 0.186 mg/L
* MnCl2.4H2O: 0.415 mg/L
* ZnCl2: 0.00327 mg/L
* FeCl3.6H2O: 0.160 mg/L
* CoCl2.6H2O: 0.00143 mg/L
* Na2MoO4.2H2O: 0.00726 mg/L
* CuCl2.2H2O: 0.000012 mg/L
* Na2EDTA.2H2O: 0.30 mg/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The culture medium was prepared using reverse osmosis purified deionised water.
- Culture medium different from test medium: no
- Intervals of water quality measurement: The pH of the control and each test preparation was determined at initiation of the test and after 72 hours exposure. The temperature within the incubator was recorded daily. The appearance of the test media was recorded daily.

OTHER TEST CONDITIONS
- Sterile test conditions: no information
- Adjustment of pH: yes, to 7.5 with 0.1 N NaOH or HCl
- Photoperiod: continuous illumination
- Light intensity and quality: intensity approximately 7000 lux provided by warm white lighting (380 to 730 nm)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
- Determination of cell concentrations [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter]: At the start of the test and after 72 h a sample of each test and control culture was removed and the cell density determined using a Coulter® Multisizer Particle Counter.
- Calculated effect parameters: specific growth rate and yield

TEST CONCENTRATIONS
- Spacing factor for test concentrations: none (limit test)
- Range finding study: yes
- Test concentrations in range finding study: 0.1, 1.0, 10 and 100% of the SS
- Results used to determine the conditions for the definitive study: yes (no significant growth inhibition was observed during the range finding study).

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes
- Any stimulation of growth found in any treatment: no
- No significant inhibition of growth rate was observed in the 100% v/v saturated solution, whereas a significant inhibition of yield by 23% was observed, compared to the control.
- 72-h EC50 (yield) > 100 mg/L
- 72-h NOEC (yield) < 100 mg/L

Results with reference substance (positive control):

- Results with reference substance valid? yes
- 72-h EC50 (growth rate) = 1.6 mg/L (95% C.L. = 1.4 to 1.8 mg/L)
- 72-h EC50 (yield) = 0.77 mg/L (95% C.L. = 0.68 to 0.87 mg/L)

Reported statistics and error estimates:

A Student’s t-test incorporating Bartlett's test for homogeneity of variance (Sokal and Rohlf, 1981) was carried out on the growth rate and yield data after 72 hours for the control and the 100% v/v saturated solution test concentration to determine any statistically significant differences between the test and control groups. All statistical analyses were performed using the SAS computer software package (SAS, 1999 - 2001).

Validity criteria fulfilled:

yes

Conclusions:

A 72-h growth inhibition test with the unicellular green alga Pseudokirchneriella subcapitata was performed with the test substance silicon zirconium oxide according to OECD guideline 201 (GLP conditions). The algae were exposed to a 100% v/v saturated solution with a nominal loading rate of 100 mg/L. No significant growth inhibition (growth rate-based) was observed in the definitive test in this saturated solution. The 72-h EC50 was therefore determined to be > 100 mg/L.

Description of key information

The study of Vryenhoef (2018), investigating the toxicity of silicon zirconium oxide to algae according to OECD guideline 201, was considered reliable without restrictions (Klimisch 1) and was considered as the key study for endpoint coverage. The EC50 for growth rate inhibition (72-h ErC50) was > 100 mg/L because no inhibition of growth rate was observed in a 100% v/v saturated solution with nominal loading rate of 100 mg/L. The substance can be considered as not harmful to algae.