lambda4-cerium(4+) zirconium(4+) tetraoxidandiide

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

from 08-NOV-2005 to 24-JAN-2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Details on sampling:

No analytical monitoring

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test item was not soluble in tap water at the required concentration and no stock solution could be prepared. Therefore, 501.9 mg of the test item was weighed by means of an analytical balance and transferred to the test flask with 284 mL of tap water. The test item was mixed into the tap water by ultrasonic treatment for ten minutes and intense stirring for 24 hours at room temperature in the dark to dissolve a maximum amount of the test item and/or disperse it as homogeneously as possible. No emulsifiers or solvents were used. After the stirring period, 16 ml of synthetic wastewater and 200 ml of the activated sludge inoculum were added.
- Differential loadings: No (limit test)
- Controls: two controls containing only tap water, synthetic wastewater and inoculum were tested in parallel to the limit test concentration of the test item, under identical test conditions.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): At the limit test concentration of 1003.8 mg/L, the test item was unevenly suspended in the turbid test medium after the 24-hour stirring period. Thus, the test concentration was clearly above the water solubility limit of cerium dioxide under the test conditions.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Laboratory culture: no, aerobic activated sludge came from a wastewater treatment plant (ARA Ergolz II, Füllinsdorf, CH) treating predominantly domestic wastewater.
- Method of cultivation: During the holding period of three days prior to use, the sludge was fed daily with 50 mL synthetic wastewater per liter and was kept at room temperature under continuous aeration until use. The pH of the activated sludge inoculum was 7.1.
- Preparation of inoculum for exposure: The sludge was washed twice with tap water by centrifugation and the supernatant liquid phase was decanted. An aliquot of washed sludge was suspended in tap water to obtain a concentration equivalent to 4 g dry material per liter.
- Pretreatment: no
- Initial biomass concentration: 4 g dry material per liter

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Post exposure observation period:

none

Hardness:

no data

Test temperature:

the temperature measured in test media in one control was 19°C at the start and 20°C at the end of the incubation period

pH:

7.3 to 8.2

Dissolved oxygen:

7.1 to 8.6 mg/L

Salinity:

not applicable

Nominal and measured concentrations:

Nominal concentration: 1003.8 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Type: open
- Material, size, headspace, fill volume: The test was performed in 2000-mL glass beakers, filled with a 500-ml mixture containing synthetic wastewater, test medium and activated sludge.
- Aeration: During the incubation period of exactly 3 hours, all test media and the controls were continuously aerated by intense stirring on magnetic stirrers, to avoid possible foaming and/or stripping of the test item.
- No. of organisms per vessel: not applicable
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2
- Biomass loading rate: sludge concentration = 1.6 g dry material / L test medium

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water
- Total organic carbon, Particulate matter, Metals, Pesticides, Chlorine, Alkalinity, Ca/mg ratio, Conductivity: no data
- Culture medium different from test medium: no
- Intervals of water quality measurement: The pH values and dissolved oxygen concentrations were determined after the addition of synthetic wastewater and activated sludge, in all test media and the controls at the start and the end of the 3-hour incubation period. The water temperature was measured in one control at the start and at the end of the incubation period. Before the addition of activated sludge and synthetic wastewater, the appearance of the test media was recorded.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Lighting: no data

EFFECT PARAMETERS MEASURED : For measurement of the respiration rate, a well-mixed sample of each test medium was poured into a BOD-flask after exactly three hours incubation time, and was not further aerated. Then, the dissolved oxygen concentration was measured with an oxygen electrode (WTW TriOxmatic 300 and an oxygen meter WTW Oxi 539, Wissenschaftlich-Technische Werkstaetten WTW, Weilheim/Germany), and was continuously recorded for about ten to fifteen minutes. During measurement, the samples were continuously stirred on a magnetic stirrer. The oxygen consumption rate (in mg O2 L-1 minute-1) was determined from the linear part of the respiration curve in the range of 6.5-2.5 mg O2/L. I n case of very rapid oxygen consumption, the range used was below the limits indicated above, but always within the linear part of the respiration curve. In case of low oxygen consumption, the rate was determined over a period of at least ten minutes.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: not applicable (limit test)
- Justification for using less concentrations than requested by guideline: a limit test has been performed to demonstrate that the test item has no toxic effect on activated sludge up to the nominal concentration of 1000 mg/L.
- Range finding study: none

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol tested at 5, 16 and 50 mg/L

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

>= 1 003.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 003.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: At the limit test concentration of 1003.8 mg/L, the test item was unevenly suspended in the turbid test medium after the 24-hour stirring period. Thus, the test concentration was clearly above the water solubility limit of cerium dioxide under the test conditions. As no adverse effect was observed at this concentration, cerium dioxide appears as non toxic for micro-organisms up to its solubility limit into water.
- Effect concentrations exceeding solubility of substance in test medium: No

Results with reference substance (positive control):

The 3-hour EC50 of the reference item 3,5-dichlorophenol (positive control) was calculated to be 12 mg/L with 95% confidence limits from 6 to 20 mg/L . The 3-hour EC50 is within the guideline-recommended range of 5 - 30 mg/L, confirming suitability of the activated sludge used.

Reported statistics and error estimates:

The 3h-EC50, -EC20 and -EC80 of the test item and their 95% confidence limits could not be calculated due to the absence of a toxic effect. For the reference substance, the probit analysis was used (slope b: 2.059, intercept a: 2.782).

Table 1: Influence of opaline de polissage (cerium oxide) and 3, 5-dichlorophenol on the oxygen consumption of activated sludge

Vessel No.	Test chemical	Nominal concentration of test chemical (mg/L)	Oxygen consumption rate (mg O2/L min-1)	Inhibition (%)
1 11	Control Control	0 0	0.973 0.978
Mean deviation (%)			0.975 0.6
2 3 4 5	3,5-dichlorophenol 3,5-dichlorophenol 3,5-dichlorophenol cerium dioxide	5 16 50 1003.8	0.756 0.402 0.092 1.046	22.5 58.8 90.6 -7.2

-% inhibition: increased oxygen consumption rate relative to control

Validity criteria fulfilled:

yes

Remarks:

The EC50 of the reference substance is in the acceptance range and the oxygen consumption rates of the 2 controls differed by less than 15% (0.6%).

Conclusions:

Cerium dioxide has no toxic effect on the respiration rate of activated sludge microorganisms at the limit test concentration of 1003.8 mg/L.

Executive summary:

The inhibitory effect of cerium dioxide on the respiration rate of aerobic wastewater microorganisms of activated sludge was investigated in a 3-hour respiration inhibition test according to the EU Commission Directive 88/302/EEC, Part C.11, and the OECD Guideline for Testing of Chemicals, No. 209.

A limit test was performed with one concentration of cerium dioxide of nominally 1003.8 mg/L.

In addition, two controls and three different concentrations of the reference item 3,5 dichlorophenol (5, 16, and 50 mg/L) were tested in parallel. After a 3-hour incubation time, the inhibition of the respiration rates was investigated via a determination of oxygen consumption rates.

The results of the reference treatments (EC50 = 12 mg/L within the guideline-recommended range of 5 - 30 mg/L) confirmed suitability of the activated sludge and the method used.

The test item cerium dioxide had no significant inhibitory effect on the respiration rate of activated sludge after the incubation period of three hours at the limit test concentration of 1003.8 mg/L. Thus, the 3-hour NOEC of cerium dioxide to activated sludge microorganisms was > = 1003.8 mg/L. The 3-hour EC20, EC50 and EC80 were clearly higher than 1003.8 mg/L.

Reference 2

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study without detailed documentation

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

not specified

Analytical monitoring:

no

Details on sampling:

none

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test material was dispersed directly in water. This approach allowed for assessment of any effects of excess undissolved test material on the activated sludge micro-organisms exoenzymes and also allowed for an evaluation of the uptake of undissolved test material by processes such as phagocytosis which may adversely affect activated sewage sludge. Additionally this procedure allowed for an examination of the entire waste water treatment facility with regards to possible initial contact toxicity.
- Controls: yes, test water without test item
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no data

Test organisms (species):

other: activated sewage sludge bacteria

Details on inoculum:

No data

Test type:

not specified

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Post exposure observation period:

None

Hardness:

No data

Test temperature:

No data

pH:

No data

Dissolved oxygen:

No data

Salinity:

No data

Nominal and measured concentrations:

Nominal concentration: 1000 mg/L cerium oxide

Details on test conditions:

TEST SYSTEM
- Test vessel: no data
- Aeration: no data
- No. of organisms per vessel: not applicable
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 2
- No. of vessels per positive control (replicates): 1
- Biomass loading rate: sludge concentration = no data

TEST MEDIUM / WATER PARAMETERS
No data

EFFECT PARAMETERS MEASURED : The oxygen consumption rate (in mg O2 L-1 minute-1) and percentage inhibition was determined.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: not applicable (limit test)
- Justification for using less concentrations than requested by guideline: a limit test has been performed to demonstrate that the test item has no toxic effect on activated sludge up to the nominal concentration of 1000 mg/L.
- Range finding study: none

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol tested at 3.2, 10 and 32 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

There was no effect on the respiration of activated sewage sludge following a 3-hour exposure to the test material at 1000 mg/L.

Results with reference substance (positive control):

- Results with reference substance valid? yes
- The 48h-EC50 was not calculated in the publication. It should be noticed that 29% inhibition was observed at 3.2 mg/L, 56% inhibition was observed at 10 mg/L, and 80% inhibition was observed at the highest tested (32.0 mg/L)

Reported statistics and error estimates:

No data

Table 1: Oxygen consumption rates and percentage inhibition values in the definitive ASRIT test after 3-h contact time

Nominal concentration (mg/L)	Initial O2 reading (mg O2/L)	Measurement period (minutes)	Final O2 reading (mg O2/L)	O2 consumption rates (mg O2/L/min)	Inhibition (%)
Control 1 2	6.9 6.3	10 8	2.8 3.0	0.41 0.41	- -
Cerium oxide 1000 1000 1000	6.7 6.7 6.8	9 10 10	3.0 2.6 2.8	0.41 0.41 0.40	0 0 2
3,5-dichlorophenol 3.2 10 32	7.1 7.7 8.2	10 10 10	4.2 6.0 7.5	0.29 0.17 0.07	29 59 83

Validity criteria fulfilled:

not specified

Conclusions:

Cerium oxide has no toxic effect on the respiration rate of activated sludge microorganisms at the limit test concentration of 1000 mg/L.

Executive summary:

The inhibitory effect of cerium oxide on the respiration rate of activated sludge microorganisms was investigated in a 3-hour respiration inhibition test according to the OECD Guideline for Testing of Chemicals, No. 209.

A limit test was performed with one concentration of cerium oxide of nominally 1000 mg/L.

In addition, two controls and three different concentrations of the reference item 3,5 dichlorophenol (3.2, 10, and 32 mg/L) were tested in parallel. After a 3-hour incubation time, the inhibition of the respiration rates were investigated via determination of oxygen consumption rates.

The test item cerium oxide had no significant inhibitory effect on the respiration rate of activated sludge after the incubation period of three hours at the limit test concentration of 1000 mg/L. Thus, the 3-hour NOEC of cerium oxide to activated sludge microorganisms was = 1000 mg/L. The 3-hour EC50 was greater than 1000 mg/L.

Reference 3

Endpoint:

activated sludge respiration inhibition testing

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

supporting study

Justification for type of information:

Read across from studies performed with micrometric and nanometric cerium dioxide. The read across justification document is attached in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Remarks on result:

other: Based on the results from studies conducted with cerium dioxide (both micrometric and nanometric forms), it can be concluded that no effects are expected for the reaction mass of cerium dioxide and zirconium dioxide.

Reference 4

Endpoint:

activated sludge respiration inhibition testing

Data waiving:

other justification

Justification for data waiving:

other:

Reference 5

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The study was conducted according to the OECD internationally recognised guideline, but published data was described with a limited level of details.

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

not specified

Analytical monitoring:

no

Details on sampling:

No analytical monitoring of the test substance concentration during the test

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test material was dispersed directly in water. This approach allowed for assessment of any effects of excess undissolved test material on the activated sludge micro-organisms exoenzymes and also allowed for an evaluation of the uptake of undissolved test material by processes such as phagocytosis which may adversely affect activated sewage sludge. Additionally this procedure allowed for an examination of the entire waste water treatment facility with regards to possible initial contact toxicity.
- Controls: yes (test water without test item)
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no data

Test organisms (species):

other: activated sewage sludge bacteria

Details on inoculum:

No data

Test type:

not specified

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Post exposure observation period:

None

Hardness:

No data

Test temperature:

No data

pH:

No data

Dissolved oxygen:

No data

Salinity:

Not applicable

Nominal and measured concentrations:

Nominal concentration: 1000 mg/L nanoparticulate cerium oxide

Details on test conditions:

TEST SYSTEM
- Test vessel: no data
- Aeration: no data
- No. of organisms per vessel: not applicable
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 2
- No. of vessels per positive control (replicates): 1
- Biomass loading rate: sludge concentration = no data

TEST MEDIUM / WATER PARAMETERS
No data

EFFECT PARAMETERS MEASURED : The oxygen consumption rate (in mg O2 L-1 minute-1) and percentage inhibition was determined.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: not applicable (limit test)
- Justification for using less concentrations than requested by guideline: a limit test has been performed to demonstrate that the test item has no toxic effect on activated sludge up to the nominal concentration of 1000 mg/L.
- Range finding study: none

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol tested at 3.2, 10 and 32 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

There was no effect on the respiration of activated sewage sludge following a 3-hour exposure to cerium oxide nanoparticles at 1000 mg/l.

Results with reference substance (positive control):

- Results with reference substance valid? yes
- The 48h-EC50 was not calculated in the publication. It should be noticed that 29% inhibition was observed at 3.2 mg/L, 56% inhibition was observed at 10 mg/L, and 80% inhibition was observed at the highest tested (32.0 mg/L) (potassium dichromate).

Table 1: Oxygen consumption rates and percentage inhibition values in the definitive ASRI test after 3h contact time

 

Nominal concentration (mg/L)	Initial O2 reading (mg O2/L)	Measurement period (minutes)	Final O2 reading (mg O2/L)	O2 consumption rates (mg O2/L/min)	% inhibition
Control 1 2	6.9 6.6	10 10	2.9 2.4	0.40 0.42	- -
nano cerium oxide 1000 1000 1000	6.7 6.8 6.5	10 10 9	2.5 2.8 2.9	0.42 0.40 0.40	[2] 2 2
3, 5-dichlorophenol 3.2 10 32	7.3 7.5 8.2	10 10 10	4.4 5.7 7.4	0.29 0.18 0.08	29 56 80

Validity criteria fulfilled:

not specified

Conclusions:

Cerium oxide nanoparticles had no toxic effect on the respiration rate of activated sludge microorganisms at the limit test concentration of 1000 mg/L.

Executive summary:

The inhibitory effect of cerium oxide nanoparticles on the respiration rate of activated sludge microorganisms was investigated in a 3-hour respiration inhibition test according to the OECD Guideline for Testing of Chemicals, n° 209.

A limit test was performed with one concentration of cerium oxide nanoparticles of nominal 1000 mg/L (3 replicates).

In addition, two controls and three different concentrations of the reference item 3,5 dichlorophenol (3.2, 10, and 32 mg/L) were tested in parallel. After a 3-hour incubation time, the inhibition of the respiration rates were investigated via determination of oxygen consumption rates.

The test item cerium oxide nanoparticles had no significant inhibitory effect on the respiration rate of activated sludge after the incubation period of three hours at the limit test concentration of 1000 mg/L. Thus, the 3-hour NOEC of cerium oxide nanoparticles to activated sludge microorganisms was = 1000 mg/L and the 3-hour EC₅₀ was greater than 1000 mg/L.

Description of key information

By analogy with one of its constituents (i.e. cerium dioxide), the reaction mass of cerium dioxide and zirconium dioxide is not expected to give rise to adverse effects in aquatic microorganisms. Therefore, no study on the reaction mass is considered necessary. 

Additional information

No data is available on the reaction mass. Based on the physicochemical, environmental and ecotoxicological data available on both the reaction mass and its two constituents cerium dioxide and zirconium dioxide, the three substances show highly similar intrinsic properties. Therefore, data available for the constituent cerium dioxide are included in the dossier as supporting information for the waiving of further studies with the reaction mass.

The following studies on cerium dioxide are included in the dossier.

Micrometric cerium dioxide: Two studies are included. One of them (Grutzner, 2006) is quoted as reliability 1 according to Klimisch and the second was given a reliability 2 (Park et al, 2007). Both studies concluded that cerium dioxide was not harmful to aquatic microoorganisms, as no adverse effects were observed at the highest dose tested (1000 mg/L).

Nanometric cerium dioxide: One literature source, scored as Klimisch 2, is available concerning the nanoparticulate form (Park et al., 2007). It concluded that nanometric cerium dioxide was not harmful to aquatic microorganisms, as no adverse effects were observed at the highest dose tested (1000 mg/L).