Ruthenium trichloride hydrate

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

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

Qualifier:

according to guideline

Guideline:

EU Method C.11 (Biodegradation: Activated Sludge Respiration Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

REPLICATES
- Positive control: 1 replicate/treatment (positive control in all experiments)
-Test item: 1 replicate/treatment 1 /10 /100 /100 mg/L (first experiment)
- Test item: 5 replicates/treatment 1000/ 460/ 220/ 100/ 46 mg/L (second and fourth experiment)
- Control: 2 replicates before and two replicates after measuring positive control and test item, respectively

CONTROLS
- 3,5-Dichlorophenol (CAS: 591-35-5) was used as a positive control. A stock solution in deiodinised water containing 500 mg/L (nominal) was freshly prepared for each experiment.
- Control: In the control vessels, 16 mL nutrient solution was mixed with 234 mL water (without ATU) and 231 mL water (with ATU), respectively.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Activated sludge: Activated sludge was taken from the activation basin of the ESN (Stadentsorgung Neustadt) sewage treatment plant in D-67435 NW-Lachen-Spreyerdorf and from the sewage treatment plant Edenkoben.
- Preparation of inoculum for exposure: Sludge was filtrated, washed with tap water three times and re-suspended in tap water.
- Pretreatment: The activated sludge was aerated until usage in the test and fed daily with 50 mL synthetic sewage feed/L.
- Initial biomass concentration:
First test: dry matter was 3.12 g suspended solids/L, giving a concentration of 1.56 g suspended solids/L in the test.
Second test: dry matter was 2.80 g suspended solids/L, giving a concentration of 1.40 g suspended solids/L in the test.
Fourth test: dry matter was 2.80 g suspended solids/L, giving a concentration of 1.40 g suspended solids/L in the test.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3

Hardness:

Analysis of drinking water hardness: 1.03 mmol/L

Test temperature:

19.1 - 20.0 °C (first experiment)
19.0 - 20.4 °C (second experiment)
18.8 - 20.9 °C (fourth experiment)

pH:

First test: 3.2 – 8.0
Second test: 3.1 – 8.1
Fourth test: 2.8 – 7.8

Nominal and measured concentrations:

First test: 1, 10, 100, 1000 mg/L
Second test: 46, 100, 220, 460, 1000 mg/L
Fourth test: 46, 100, 220, 460, 1000 mg/L
Test solutions not analysed, nominal concentrations only

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beaker
- Duration: 3 hours
- Water: tap water
- Aeration: purified air, using Pasteur pipettes
- Feeding: nutrient solution, 16 mL/vessel

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water
- Total organic carbon (TOC): 0.5 mg/L
- Pesticides and biocides: - Chloride: 12 mg/L
- Chlorinated organic compounds: - Conductivity at 25°C: 242 µS/cm

Reference substance (positive control):

yes

Remarks:

3,5-Dichlorophenol (CAS-No. 591-35-5)

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

220 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Remarks:

Inhibition based on total respiration without ATU

Remarks on result:

other: --

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

360 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Remarks:

based on total respiration without ATU

Remarks on result:

other: 95% Confidence Interval 310 - 400 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

530 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Remarks:

based on total respiration without ATU

Remarks on result:

other: 95% Confidence Interval 480 - 580 mg/L

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

220 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks:

with ATU

Remarks on result:

other: --

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

230 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks:

with ATU

Remarks on result:

other: 95% Confidence Interval 230 - 240 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

330 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks:

with ATU

Remarks on result:

other: 95% Confidence Interval 320 - 340 mg/L

Results with reference substance (positive control):

3,5-Dichlorophenol was used as positive control. Four concentrations, with and without ATU, were tested. The 3-hour EC50s of the positive were determined as 14 - 15 mg/L with N-allylthiourea (ATU) and 9.5 - 11 mg/L without ATU. The values were within the recommended range.

Reported statistics and error estimates:

CALCULATIONS
- O2 Consumption: The O2 consumption rate was calculated from the slope of the linear part of the O2 consumption curve using linear regression.
- Calculation of EC50: The estimation of EC50 was accomplished using the software Origin TM 9.0.

Validity criteria fulfilled:

yes

Remarks:

Positive control EC50s were in the required range, the COV of oxygen uptake rate was <30% in the control, oxygen uptake rate in control met the required criteria except for one value from the rangefinder which is not considered critical

Conclusions:

The 3-h EC50 was 530 mg L-1 (without ATU) and 330 mg L-1 (with ATU). The EC10 was 360 mg L-1 (without ATU) and 230 mg L-1 (with ATU). The NOEC was 220 mg L-1 both with and without ATU.

Executive summary:

An activated sludge respiration inhibition test following OECD guideline 209 was conducted (Muckle 2016). The respiration rates of samples of activated sludge fed with synthetic sewage were measured in an enclosed cell containing an oxygen electrode after a contact time of 3 hours. The sensitivity of each batch of activated sludge was checked with a positive control (3,5-Dichlorophenol); all values were within the recommended range for the positive control.

In the first experiment, which was performed as the range finding test, the test item was tested using 4 concentrations ranging from 1 to 1000 mg L-1 nominal concentration. Two test series using the same concentration were performed in order to discern between inhibition of nitrifiers and inhibition of the total population of heterotrophic bacteria. Therefore, the nitrification inhibitor N-allylthiourea (ATU) was added to all flasks for the second test series.

In the second experiment the test item was assessed using 5 concentrations ranging from 46 to 1000 mg L-1 nominal concentration without ATU. The dry matter of the inoculum was determined as 2.80 g suspended solids/L, giving a concentration of 1.40 g suspended solids in the test. The fourth experiment was conducted with the same concentrations but with ATU.

The 3-h EC50 was 530 mg L-1 (without ATU) and 330 mg L-1 (with ATU). The EC10 was 360 mg L-1 (without ATU) and 230 mg L-1 (with ATU). The NOEC was 220 mg L-1 both with and without ATU.

Description of key information

The 3-h EC50 was 530 mg L-1 (without ATU) and 330 mg L-1 (with ATU). The EC10 was 360 mg L-1 (without ATU) and 230 mg L-1 (with ATU). The NOEC was 220 mg L-1 both with and without ATU.

Additional information

An activated sludge respiration inhibition test following OECD guideline 209 was conducted (Muckle 2016). The respiration rates of samples of activated sludge fed with synthetic sewage were measured in an enclosed cell containing an oxygen electrode after a contact time of 3 hours. The sensitivity of each batch of activated sludge was checked with a positive control (3,5-Dichlorophenol); all values were within the recommended range for the positive control.

In the first experiment, which was performed as the range finding test, the test item was tested using 4 concentrations ranging from 1 to 1000 mg L-1 nominal concentration. Two test series using the same concentration were performed in order to discern between inhibition of nitrifiers and inhibition of the total population of heterotrophic bacteria. Therefore, the nitrification inhibitor N-allylthiourea (ATU) was added to all flasks for the second test series.

In the second experiment the test item was assessed using 5 concentrations ranging from 46 to 1000 mg L-1 nominal concentration without ATU. The dry matter of the inoculum was determined as 2.80 g suspended solids/L, giving a concentration of 1.40 g suspended solids in the test. The fourth experiment was conducted with the same concentrations but with ATU.

The 3-h EC50 was 530 mg L-1 (without ATU) and 330 mg L-1 (with ATU). The EC10 was 360 mg L-1 (without ATU) and 230 mg L-1 (with ATU). The NOEC was 220 mg L-1 both with and without ATU.