Dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (1:2)

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:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-compliant, guideline study, available as an unpublished report, fully adequate for assessment, reliable without restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

yes

Remarks:

Few deviations occurred: -The measurement period in the experiments was partly less than 5 minutes. -The concentration of suspended solids in the 1st Exp. was higher than stated. -Both deviations were stated to be uncritical.

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

Few deviations occurred: -The measurement period in the experiments was partly less than 5 minutes. -The concentration of suspended solids in the 1st Exp. was higher than stated. -Both deviations were stated to be uncritical.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION:
- Method: due to the poor solubility of the test item, the test item was weighed directly into the test vessels. Afterwards, 16 mL nutrient solution and water were added to give 250 mL. Then, 250 mL inoculum was added in 5-minute intervals and the mixtures were aerated
- Controls: a blank control was prepared by mixing 16 mL nutrient solution with 234 mL water (without ATU) and 231 mL water (with ATU), respectively
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): not reported

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Name and location of sewage treatment plant where inoculum was collected: the sludge was taken from the activation basin of sewage treatment plant in D-67480 Edenkoben
- Preparation of inoculum for exposure: Sludge was filtrated, washed with tap water three times and re-suspended in tap water. The activated sludge was aerated until usage in the test and fed daily with 50 mL synthetic sewage feed/L.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Hardness:

- Hardness: 1.03 mmol/L

Test temperature:

First experiment: 19.4 - 21.0 °C
Second experiment: 19.1 - 21.5 °C
Third experiment: 19.6 - 21.3 °C

pH:

First experiment: 8.2 - 8.3
Second experiment: 7.9 - 8.1
Third experiment: 7.9 - 8.0

Nominal and measured concentrations:

Nominal concentrations: 1, 10, 100, 1000 mg test item/L (first experiment); 60, 130, 290, 640, 1400 mg test item/L (second and third experiments)

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers as test vessels; narrow-neck glass bottles with flat bottoms (250 mL) as measuring flasks
- Aeration: purified air, using Pasteur pipettes
- No. of vessels per concentration (replicates): 1 (first experiment), 5 (second and third experiments)
- No. of vessels per control (replicates): 2 before and 2 after measuring positive control and test item, respectively
- No. of vessels per positive control (replicates): 1
- Sludge concentration (weight of dry solids per volume): First experiment: the dry matter was determined as 3.36 g suspended solids, giving a concentration of 1.68 g suspended solids/L in the test; second experiment: the dry matter was determined as 2.80 g suspended solids, giving a concentration of 1.40 g suspended solids/L in the test; third experiment: the dry matter was determined as 2.80 g suspended solids, giving a concentration of 1.40 g suspended solids/L in the test
- Nutrients provided for bacteria: synthetic sewage used as nutrient solution, containing peptone 16.0 g, meat extract 11.0 g, urea 3.0 g, NaCl 0.7 g, CaCl2.2H2O 0.4 g, MgSO4.7H2O 0.2 g, K2HPO4 2.8 g and deionised water ad 1000 mL.
- Nitrification inhibitor used: N-allylthiourea (ATU)
- Biomass loading rate: 250 mL (as inoculum)

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water (Total organic carbon (TOC) 0.5 mg/L, Pesticides and biocides < LOD, Chloride 12 mg/L, Chlorinated organic compounds < LOD, Conductivity 242 µS/cm at 25 °C, Hardness 1.03 mmol/L)

OTHER TEST CONDITIONS
- Photoperiod: Not reported
- Light intensity: Not reported

EFFECT PARAMETERS MEASURED: the respiration rate was determined by measurement of the oxygen concentration over a period of maximum 5 minutes after 3 hours contact time

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10 (First experiment), about 2.2 (second and third experiments)
- Range finding study (First experiment)
- Test concentrations: 1, 10, 100 and 1000 mg test item/L
- Results used to determine the conditions for the definitive study: significant inhibition was observed

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Key result

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

60 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other: Without ATU.

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

10.29 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Remarks on result:

other: Without ATU.

Key result

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

120 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 84- 150 mg/L. Without ATU.

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

20.58 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 14.41 - 25.73 mg/L. Without ATU.

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

1 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 850 - 1400 mg/L. Without ATU.

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

188.65 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 146 - 240.1 mg/L. Without ATU.

Key result

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

130 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: With ATU.

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

22.3 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: With ATU.

Key result

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

170 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 140 - 210 mg/L. With ATU.

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

29.16 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 24.01 - 36.02 mg/L. With ATU.

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

1 400 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: not determined. With ATU.

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

240.1 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: not determined. With ATU.

Details on results:

The oxygen consumption rate was calculated from the slope of the linear part of the O2 consumption curve using linear regression.

Results with reference substance (positive control):

Four concentrations of the positive control, with and without ATU, were tested. Three experiments were performed as well. In the first experiment, the 3-hour EC50 values of the positive were determined as 15 mg/L with N-allylthiourea (ATU) and 13 mg/L without ATU. In the second experiment, the 3-h EC50 without ATU was determined to be 11 mg/L, and in the third experiment the 3-h EC50 could not be determined.
All values were within the recommended range of 2 - 25 mg/L (total respiration without ATU), and of 5 - 40 mg/L (heterotrophic respiration with ATU)

Reported statistics and error estimates:

EC50 values were determined using linear fit on a probability-logarithmic scale.

Table 1. Oxygen consumption and % Inhibition after 3 hours contact time to dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (First experiment without ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	42.916	0
Control	0	42.512	0
Control	0	43.972	0
Control	0	43.410	0
Control	0	42.888	0
Control	0	43.432	0
Test item	1	43.434	-1.0
Test item	10	43.168	-0.3
Test item	100	38.047	11.6
Test item	1000	22.519	47.7
Positive Control	5	36.426	15.3
Positive Control	10	24.775	42.4
Positive Control	20	11.940	72.2
Positive Control	40	6.664	84.5

Table 2. Oxygen consumption and % Inhibition after 3 hours contact time to dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (First experiment with ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	42.500	0
Control	0	40.921	0
Control	0	44.149	0
Control	0	38.279	0
Control	0	42.149	0
Control	0	41.327	0
Test item	1	43.626	-5.0
Test item	10	41.057	1.2
Test item	100	38.358	7.7
Test item	1000	23.018	44.6
Positive Control	5	38.828	6.6
Positive Control	10	26.355	36.6
Positive Control	20	11.823	71.5
Positive Control	40	6.730	83.8

Table 3. Oxygen consumption and % Inhibition after 3 hours contact time to dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (Second experiment without ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	34.127	0
Control	0	37.392	0
Control	0	34.512	0
Control	0	38.847	0
Control	0	35.689	0
Control	0	35.960	0
Test item	60	36.786	-1.9
Test item	60	36.643	-1.5
Test item	60	36.214	-0.3
Test item	60	35.562	1.5
Test item	60	37.526	-4.0
Test item	130	31.580	12.5
Test item	130	31.816	11.8
Test item	130	30.574	15.3
Test item	130	32.357	10.3
Test item	130	34.386	4.7
Test item	290	25.583	29.1
Test item	290	27.589	23.6
Test item	290	26.111	27.6
Test item	290	24.581	31.9
Test item	290	26.574	26.4
Test item	640	20.140	44.2
Test item	640	18.864	47.7
Test item	640	20.706	42.6
Test item	640	20.126	44.2
Test item	640	20.825	42.3
Test item	1400	17.47	51.6
Test item	1400	21.031	41.7
Test item	1400	18.277	49.4
Test item	1400	18.369	49.1
Test item	1400	17.997	50.1
Positive control	5	29.789	17.5
Positive control	10	16.639	53.9
Positive control	20	7.758	78.5
Positive control	40	4.379	87.9

Table 4. Oxygen consumption and % Inhibition after 3 hours contact time to dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (Third experiment with ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	25.490	0
Control	0	28.872	0
Control	0	28.844	0
Control	0	25.880	0
Control	0	31.555	0
Control	0	28.993	0
Test item	60	27.324	3.4
Test item	60	29.443	-4.1
Test item	60	28.545	-1.0
Test item	60	27.175	3.9
Test item	60	27.182	3.9
Test item	130	26.167	7.4
Test item	130	27.553	2.5
Test item	130	27.310	3.4
Test item	130	29.239	-3.4
Test item	130	25.004	11.6
Test item	290	24.515	13.3
Test item	290	22.981	18.7
Test item	290	25.228	10.8
Test item	290	23.388	17.3
Test item	290	25.544	9.6
Test item	640	16.835	40.5
Test item	640	16.174	42.8
Test item	640	16.590	41.3
Test item	640	16.640	41.1
Test item	640	16.995	39.9
Test item	1400	14.901	47.3
Test item	1400	15.238	46.1
Test item	1400	15.239	46.1
Test item	1400	15.075	46.7
Test item	1400	14.837	47.5
Positive control	5	27.7	2.0
Positive control	10	16.353	42.2
Positive control	20	7.757	72.6
Positive control	40	4.553	83.9

Validity criteria fulfilled:

yes

Conclusions:

Based on nominal concentrations, the 3-h EC50, EC10 and NOEC without ATU for dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, were determined to be 1100 mg test item/L (188.65 mg Pt/L), 120 mg test item/L (20.58 mg Pt/L), and 60 mg test item/L (10.29 mg Pt/L), respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 1400 mg test item/L (240.1 mg Pt/L), 170 mg test item/L(29.16 mg Pt/L), and 130 mg test item/L(22.3 mg Pt/L), respectively.

Executive summary:

An activated sludge respiration inhibition test was conducted with dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol. The study is reliable without restrictions, being GLP-compliant and having followed standard test guidelines (OECD 209 and EU Method C.11).

Two test series using the same concentrations were carried out, with and without the nitrification inhibitor N- allylthiourea (ATU) to discern between inhibition of nitrificators and inhibition of the total population. Because significant inhibition was observed, two additional experiments with and without ATU, respectively, were performed under the same test conditions. A positive control was used, and all validity criteria were met. Based on nominal concentrations, the 3-h EC50, EC10 and NOEC without ATU for dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, were determined to be 1100 mg test item/L (188.65 mg Pt/L), 120 mg test item/L (20.58 mg Pt/L), and 60 mg test item/L (10.29 mg Pt/L), respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 1400 mg test item/L (240.1 mg Pt/L), 170 mg test item/L(29.16 mg Pt/L), and 130 mg test item/L(22.3 mg Pt/L), respectively.

Reference 2

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:

other: GLP-compliant, guideline study, available as an unpublished report, fully adequate for assessment, reliable without restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

yes

Remarks:

Few deviations occurred (i.e. T°C and measurement period differed from the study plan). Both reported to be uncritical, normal respiration activity of the control was observed and correlation of the inhibition values within the test replicates was good.

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

Few deviations occurred (i.e. T°C and measurement period differed from the study plan). Both reported to be uncritical, normal respiration activity of the control was observed and correlation of the inhibition values within the test replicates was good.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Due to the poor solubility of the test item, the test item was weighed directly into the test vessels.
In the control vessels, 16 mL nutrient solution was mixed with 234 mL water (without N-allylthiourea (ATU)), and with 231 mL water (with ATU), respectively. Both the positive control vessels and the treatments were prepared by putting the appropriate amount of positive control solution or test item, with the addition of ATU into the respective test vessel (the concentration of the positive control was calculated using the concentration of the stock solution and the dilution factor. The test item was added to the test vessels directly). Then, 16 mL nutrient solution and water were added to give 250 mL. Lastly, 250 mL inoculum was added in 5 minute intervals to all vessels, and the mixtures were aerated.
After 3 hours, the content of the first vessel was poured in a 250 mL narrow-neck bottle and the respiration rate was determined by measurement of the oxygen-concentration over a period of max 5 minutes. The following vessels were measured in 5 minute intervals.
- Differential loading: Nominal concentrations of the test item were 1, 10, 100, 1000 mg/L (first experiment, range finder), and 1, 3.2, 10, 32, 100 and 320 mg/L (second and third experiment).
- Controls: both blank and positive controls were used. Blank controls contained an equal volume of activated sludge, tap water and nutrient solution. 3,5-Dichlorophenol (CAS n. 591-35-5) was used as positive control, and a stock solution in deionised water containing 500 mg/L (nominal) was freshly prepared for each experiment.
- Nutrient solution: synthetic sewage.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): None

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Name and location of sewage treatment plant where inoculum was collected: acitivation basin of the ESN (Stadtentsorgung Neustadt) sewage treatment plant D-67435 NW-Lachen-Speyerdorf
- Preparation of inoculum for exposure: On the day before the experiment, the inoculum was taken from its source, filtrated, washed with tap water and resuspended in tap water. The activated sludge was aerated until usage in the test and the dry matter was determined. The sludge was fed daily with 50 mL sludge/L.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Hardness:

-

Test temperature:

First experiment: 22.2 - 23.1 °C
Second experiment: 19.5 - 21.5 °C
Third experiment: 21.0 - 22.9 °C

pH:

First experiment: 6.4 - 8.1
Second experiment: 7.7 - 8.1
Thrid experiment: 8.0 - 8.2

Dissolved oxygen:

-

Salinity:

-

Nominal and measured concentrations:

Nominal concentrations: 1, 10, 100 and 1000 mg test item/L (first experiment); 1, 3.2, 10, 32, 100 and 320 mg test item/L (second and third experiments)

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers as test vessels. Narrow-neck glass bottles with flat bottoms (250 mL) as measuring flasks
- Aeration: Purified air, using Pasteur pipettes
- Renewal rate of test solution (frequency/flow rate): No renewal
- No. of vessels per concentration (replicates): 1 (first experiment), 5 (second and third experiments)
- No. of vessels per control (replicates): 6
- No. of vessels per positive control (replicates): 1
- Sludge concentration (weight of dry solids per volume): Exp. 1) the dry matter of the inoculum was determined as 2.94 g suspended solids/L, giving a concentration of 1.47 g suspended solids/L in the test; Exp. 2) the dry matter of the inoculum was determined as 3.14 g suspended solids/L, giving a concentration of 1.57 g suspended solids/L in the test; Exp. 3) the dry matter of the inoculum was determined as 2.42 g suspended solids/L, giving a concentration of 1.21 g suspended solids/L in the test.
- Nutrients provided for bacteria: synthetic sewage containing peptone 16.0 g, meat extract 11.0 g, urea 3.0 g, NaCl 0.7 g, CaCl2.2H2O 0.4 g, MgSO4.7H2O 0.2 g, K2HPO4 2.8 g and deionised water ad 1000 mL. The pH of the solution was 7.1. After preparation, the nutrient solution was frozen immediately.
- Nitrification inhibitor used: N-allylthiourea (first and third experiments)
- Biomass loading rate: 250 mL inoculum per treatment

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water (pH 8.20, Total Organic Carbon 0.24 mg/L, Hardness 1.10 mmol/L)

OTHER TEST CONDITIONS
- Photoperiod: not reported
- Light intensity: not reported

EFFECT PARAMETERS MEASURED: the respiration rate was determined by measurement of the oxygen concentration over a period of maximum 5 minutes after 3 hours of contact time.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10 (first experiment) and 3.2 (second and third experiment)
- Range finding study
- Test concentrations: 1, 10, 100, 1000 mg test item/L (nominal concentrations - first experiment)
- Results used to determine the conditions for the definitive study: no significant inhibition was observed

Reference substance (positive control):

yes

Remarks:

3,5-Dichlorophenol (CAS n. 591-35-5)

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

3.2 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.:

1.26 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 5.5 - 6.5 mg/L

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

2.35 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 2.15 - 2.55 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

103 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 96 - 110 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

40.33 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 37.58 - 43.065 mg/L

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

3.92 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

8.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 6.0 - 11 mg/L

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

3.21 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 2.35 - 4.31 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

83 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 76 - 91 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

32.49 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 29.75 - 35.63 mg/L

Details on results:

- The oxygen consumption rate was calculated from the slope of the linear part of the oxygen consumption curve using linear regression.
- The inihibition values in the three experiments (with and without ATU) showed good correlation of the inhibitory effects of the test item.
- In the three highest concentration treatments of the test series with and without ATU, the inhibition of the respiration was in the same range. However, no inhibition was observed at a test item concentration of 10 mg test item/L with ATU. Whereas, significant inhibition was observed within test series treated without ATU at the concentration of 10 mg test item/L. Most likely, at a limiting concentration of 10 mg test item/L, the test item acted as nitrification inhibitor, whilst at higher concentrations inhibition of total population with and without ATU was similar.

Results with reference substance (positive control):

In the first experiment, a 3 h-EC50 value of 12 mg/L was determined with ATU, and a 3 h-EC50 value of 16 mg/L without ATU.
In the second and third experiments, 3 h-EC50 values of 12 mg/L (without ATU) and 17 mg/L (with ATU) were determined, respectively.
All values lay within the recommended ranges of 2 - 25 mg/L (total respiration without ATU), and 5 - 40 mg/L (heterotrophic respiration with ATU).

Reported statistics and error estimates:

Both NOEC values were selected after having investigated likely differences between treatments and controls, firstly using an F-test, and afterwards either a WEIR or a t-test.
For the calculation of the EC10 and EC50, the percentage of ihibition was plotted versus concentration in a Gauβ-logarithmic diagram. EC10 and EC50 were determined from the x values of the regression line at y = 10 % and y = 50 %. The data were evaluated using a linear fit on a probability-logarithmic scale.

Table 1. Oxygen consumption and % Inhibition after 3 hours contact time to hexachloroplatinic acid (First experiment without ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	75.308	0
Control	0	72.253	0
Control	0	75.341	0
Control	0	69.683	0
Control	0	74.287	0
Control	0	75.570	0
Test item	1	67.083	9.0
Test item	10	59.554	19.2
Test item	100	33.849	54.1
Test item	1000	8.609	88.3
Positive Control	5	63.626	13.7
Positive Control	10	39.725	46.1
Positive Control	20	18.156	75.4
Positive Control	40	7.572	89.7

Table 2. Oxygen consumption and % Inhibition after 3 hours contact time to hexachloroplatinic acid (First experiment with ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	66.286	0
Control	0	65.840	0
Control	0	68.037	0
Control	0	66.438	0
Control	0	62.647	0
Control	0	64.679	0
Test item	1	64.833	1.3
Test item	10	50.010	23.8
Test item	100	26.511	59.6
Test item	1000	4.886	92.6
Positive Control	5	64.313	2.0
Positive Control	10	40.337	38.6
Positive Control	20	18.990	71.1
Positive Control	40	7.782	88.1

Table 3. Oxygen consumption and % Inhibition after 3 hours contact time to hexachloroplatinic acid (Second experiment without ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	60.026	0
Control	0	58.551	0
Control	0	64.083	0
Control	0	61.690	0
Control	0	51.771	0
Control	0	55.800	0
Test item	1	62.577	-6.7
Test item	1	64.217	-9.5
Test item	1	64.113	-9.3
Test item	1	55.289	5.7
Test item	1	54.591	6.9
Test item	3.2	55.457	5.4
Test item	3.2	55.126	6.0
Test item	3.2	55.726	5.0
Test item	3.2	55.245	5.8
Test item	3.2	60.789	-3.6
Test item	10	50.130	14.5
Test item	10	49.991	14.8
Test item	10	49.060	16.4
Test item	10	50.355	14.1
Test item	10	49.054	16.4
Test item	32	38.749	33.9
Test item	32	40.500	30.9
Test item	32	41.862	28.6
Test item	32	39.587	32.5
Test item	32	39.034	33.5
Test item	100	30.235	48.5
Test item	100	31.103	47.0
Test item	100	30.074	48.7
Test item	100	27.167	53.7
Test item	100	28.749	50.9
Test item	320	18.392	68.6
Test item	320	17.853	69.6
Test item	320	18.851	67.9
Test item	320	18.580	68.3
Test item	320	19.044	67.5
Positive control	5	48.979	16.5
Positive control	10	31.034	47.1
Positive control	20	16.346	72.1
Positive control	40	8.009	86.3

Table 4. Oxygen consumption and % Inhibition after 3 hours contact time to hexachloroplatinic acid (Third experiment with ATU)

ontent	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	40.419	0
Control	0	39.976	0
Control	0	45.050	0
Control	0	43.806	0
Control	0	49.121	0
Control	0	50.868	0
Test item	1	54.645	-21.8
Test item	1	49.270	-9.8
Test item	1	53.765	-19.8
Test item	1	50.530	-12.6
Test item	1	50.762	-13.1
Test item	3.2	50.669	-12.9
Test item	3.2	54.287	-21.0
Test item	3.2	52.260	-16.5
Test item	3.2	51.606	-15.0
Test item	3.2	49.048	-9.3
Test item	10	45.370	-1.1
Test item	10	46.688	-4.0
Test item	10	46.598	-3.8
Test item	10	47.739	-6.4
Test item	10	47.693	-6.3
Test item	32	31.240	30.4
Test item	32	33.082	26.3
Test item	32	31.308	30.2
Test item	32	32.971	26.5
Test item	32	32.262	28.1
Test item	100	18.114	59.6
Test item	100	19.390	56.8
Test item	100	18.232	59.4
Test item	100	18.777	58.2
Test item	100	18.071	59.7
Test item	320	11.270	74.9
Test item	320	10.793	75.9
Test item	320	11.160	75.1
Test item	320	11.038	75.4
Test item	320	10.744	76.1
Positive control	5	41.102	8.4
Positive control	10	37.805	15.8
Positive control	20	14.931	66.7
Positive control	40	7.764	82.7

Validity criteria fulfilled:

yes

Conclusions:

The inhibition of the total respiration of activated sludge when exposed to hexachloroplatinic acid, with and without the addition of the nitrification inhibitor N-allylthiourea was assessed. Based on nominal concentrations, the 3-h EC50, EC10 and NOEC values without ATU were determined to be 103 mg test item L-1 (40.33 mg Pt L-1), 6.0 mg test item L-1 (2.35 mg Pt L-1), and 3.2 mg test item L-1 (1.26 mg Pt L-1), respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 83 mg test item L-1 (32.49 mg Pt L-1), 8.2 mg test item L-1 (3.21 mg Pt L-1), and 10 mg test item L-1 (3.92 mg Pt L-1), respectively.

Executive summary:

The inhibition of the respiration of activated sludge when exposed to the test item hexachloroplatinic acid was investigated. The study is reliable without restrictions, being GLP-compliant and having followed the standard test guidelines (OECD 209 and EU-Method C.11).

Two test series using the same concentrations were carried out, with and without the nitrification inhibitor N- allylthiourea (ATU) to discern between inhibition of nitrificators and inhibition of the total population. Because significant inhibition was observed, two additional experiments with and without ATU, respectively, were performed under the same test conditions. A positive control was used, and all validity criteria were met.

Based on nominal concentrations, the 3-h EC50, EC10 and NOEC values without ATU were determined to be 103 mg test item L^-1(40.33 mg Pt L^-1), 6.0 mg test item L^-1(2.35 mg Pt L^-1), and 3.2 mg test item L^-1(1.26 mg Pt L^-1), respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 83 mg test item L^-1(32.49 mg Pt L^-1), 8.2 mg test item L^-1(3.21 mg Pt L^-1), and 10 mg test item L^-1(3.92 mg Pt L^-1), respectively.

Reference 3

Endpoint:

activated sludge respiration inhibition testing

Type of information:

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

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP-compliant, guideline study, available as an unpublished report, fully adequate for assessment, reliable without restrictions

Justification for type of information:

1. HYPOTHESIS FOR THE ANALOGUE APPROACH

The general principles applied for read across between metal substances are that ecotoxicity and the potential for adverse environmental effects are based on the metal ion in cases where the counter ions can reasonably be expected to be non-toxic, as is the case for many simple metals salts (e.g. anions such as SO42-, NO3-, OH-).

When reading across between different metal substances, the oxidation state of the metal ion needs to be carefully considered. For metals, chemical speciation can affect both the fate of the substance in the environment and its toxicity. For some metals (e.g. chromium and arsenic), large differences in environmental toxicity between difference oxidation states have been observed. For platinum substances, the database of ecotoxicity data is not as extensive as for other metal substances, but there may be a difference in toxicity between platinum (II) and platinum (IV) substances.

For platinum (IV) substances a slightly different read across approach is adopted for fulfilment of hazard endpoints and classification, compared to the approach for risk assessment. The approach for hazard data and classification takes into account potential differences in toxicity due to the co-ordinating ligands, whereas for risk assessment a worst case approach is followed in order to enable the comparison of total measured platinum(IV) concentrations in the environment with a single PNEC derived for platinum(IV) substances.

2. SOURCE AND TARGET CHEMICAL(S)

Source chemical: Hexachloroplatinic acid
Target chemical: Dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol

3. ANALOGUE APPROACH JUSTIFICATION

For platinum substances, the database of ecotoxicity data is not as extensive as for other metal substances, but there may be a difference in toxicity between platinum (II) and platinum (IV) substances. For this reason, read-across between different substances is limited to metal compounds in which the metal exists in the same oxidation state.

The approach that is applied for deriving predicted no effect concentrations (PNECs) for platinum (IV) substances assumes that the platinum present in the environment has the potential to be in the most toxic form, hexachloroplatinic acid. The generic PNECs for all platinum (IV) substances are therefore derived based upon the ecotoxicity data for hexachloroplatinic acid.

For risk assessment purposes it is acceptable to read across toxicity data from the most toxic substance, because the risk assessment process is iterative and allows scope for further refinement at higher tiers should this be appropriate to understand the potential risks posed.

In the exposure assessment, analytical monitoring of platinum emissions is unlikely to differentiate between different platinum (IV) substances. Therefore, if PNEC values were derived separately for each platinum (IV) substance it would not be possible to practically use these values in the risk assessment, when most sites use multiple platinum substances. Since environmental monitoring does not distinguish between particular platinum substances, for the risk assessment platinum (IV) in the environment is considered to be in the most toxic form.

Reason / purpose for cross-reference:

read-across source

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

3.2 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.:

1.26 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 5.5 - 6.5 mg/L

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

2.35 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 2.15 - 2.55 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

103 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 96 - 110 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

40.33 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of total respiration

Remarks on result:

other: 95% Confidence Interval: 37.58 - 43.065 mg/L

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

3.92 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

8.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 6.0 - 11 mg/L

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

3.21 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 2.35 - 4.31 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

83 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 76 - 91 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

32.49 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Pt

Basis for effect:

inhibition of heterotrophic respiration

Remarks on result:

other: 95% Confidence Interval: 29.75 - 35.63 mg/L

Details on results:

- The oxygen consumption rate was calculated from the slope of the linear part of the oxygen consumption curve using linear regression.
- The inihibition values in the three experiments (with and without ATU) showed good correlation of the inhibitory effects of the test item.
- In the three highest concentration treatments of the test series with and without ATU, the inhibition of the respiration was in the same range. However, no inhibition was observed at a test item concentration of 10 mg test item/L with ATU. Whereas, significant inhibition was observed within test series treated without ATU at the concentration of 10 mg test item/L. Most likely, at a limiting concentration of 10 mg test item/L, the test item acted as nitrification inhibitor, whilst at higher concentrations inhibition of total population with and without ATU was similar.

Results with reference substance (positive control):

In the first experiment, a 3 h-EC50 value of 12 mg/L was determined with ATU, and a 3 h-EC50 value of 16 mg/L without ATU.
In the second and third experiments, 3 h-EC50 values of 12 mg/L (without ATU) and 17 mg/L (with ATU) were determined, respectively.
All values lay within the recommended ranges of 2 - 25 mg/L (total respiration without ATU), and 5 - 40 mg/L (heterotrophic respiration with ATU).

Table 1. Oxygen consumption and % Inhibition after 3 hours contact time to hexachloroplatinic acid (First experiment without ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	75.308	0
Control	0	72.253	0
Control	0	75.341	0
Control	0	69.683	0
Control	0	74.287	0
Control	0	75.570	0
Test item	1	67.083	9.0
Test item	10	59.554	19.2
Test item	100	33.849	54.1
Test item	1000	8.609	88.3
Positive Control	5	63.626	13.7
Positive Control	10	39.725	46.1
Positive Control	20	18.156	75.4
Positive Control	40	7.572	89.7

Table 2. Oxygen consumption and % Inhibition after 3 hours contact time to hexachloroplatinic acid (First experiment with ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	66.286	0
Control	0	65.840	0
Control	0	68.037	0
Control	0	66.438	0
Control	0	62.647	0
Control	0	64.679	0
Test item	1	64.833	1.3
Test item	10	50.010	23.8
Test item	100	26.511	59.6
Test item	1000	4.886	92.6
Positive Control	5	64.313	2.0
Positive Control	10	40.337	38.6
Positive Control	20	18.990	71.1
Positive Control	40	7.782	88.1

Table 3. Oxygen consumption and % Inhibition after 3 hours contact time to hexachloroplatinic acid (Second experiment without ATU)

Content	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	60.026	0
Control	0	58.551	0
Control	0	64.083	0
Control	0	61.690	0
Control	0	51.771	0
Control	0	55.800	0
Test item	1	62.577	-6.7
Test item	1	64.217	-9.5
Test item	1	64.113	-9.3
Test item	1	55.289	5.7
Test item	1	54.591	6.9
Test item	3.2	55.457	5.4
Test item	3.2	55.126	6.0
Test item	3.2	55.726	5.0
Test item	3.2	55.245	5.8
Test item	3.2	60.789	-3.6
Test item	10	50.130	14.5
Test item	10	49.991	14.8
Test item	10	49.060	16.4
Test item	10	50.355	14.1
Test item	10	49.054	16.4
Test item	32	38.749	33.9
Test item	32	40.500	30.9
Test item	32	41.862	28.6
Test item	32	39.587	32.5
Test item	32	39.034	33.5
Test item	100	30.235	48.5
Test item	100	31.103	47.0
Test item	100	30.074	48.7
Test item	100	27.167	53.7
Test item	100	28.749	50.9
Test item	320	18.392	68.6
Test item	320	17.853	69.6
Test item	320	18.851	67.9
Test item	320	18.580	68.3
Test item	320	19.044	67.5
Positive control	5	48.979	16.5
Positive control	10	31.034	47.1
Positive control	20	16.346	72.1
Positive control	40	8.009	86.3

Table 4. Oxygen consumption and % Inhibition after 3 hours contact time to hexachloroplatinic acid (Third experiment with ATU)

ontent	Nominal Concentration in mg/L	O2 consumption in mg/(L*h)	% Inhibition
Control	0	40.419	0
Control	0	39.976	0
Control	0	45.050	0
Control	0	43.806	0
Control	0	49.121	0
Control	0	50.868	0
Test item	1	54.645	-21.8
Test item	1	49.270	-9.8
Test item	1	53.765	-19.8
Test item	1	50.530	-12.6
Test item	1	50.762	-13.1
Test item	3.2	50.669	-12.9
Test item	3.2	54.287	-21.0
Test item	3.2	52.260	-16.5
Test item	3.2	51.606	-15.0
Test item	3.2	49.048	-9.3
Test item	10	45.370	-1.1
Test item	10	46.688	-4.0
Test item	10	46.598	-3.8
Test item	10	47.739	-6.4
Test item	10	47.693	-6.3
Test item	32	31.240	30.4
Test item	32	33.082	26.3
Test item	32	31.308	30.2
Test item	32	32.971	26.5
Test item	32	32.262	28.1
Test item	100	18.114	59.6
Test item	100	19.390	56.8
Test item	100	18.232	59.4
Test item	100	18.777	58.2
Test item	100	18.071	59.7
Test item	320	11.270	74.9
Test item	320	10.793	75.9
Test item	320	11.160	75.1
Test item	320	11.038	75.4
Test item	320	10.744	76.1
Positive control	5	41.102	8.4
Positive control	10	37.805	15.8
Positive control	20	14.931	66.7
Positive control	40	7.764	82.7

Validity criteria fulfilled:

yes

Conclusions:

The inhibition of the total respiration of activated sludge when exposed to hexachloroplatinic acid, with and without the addition of the nitrification inhibitor N-allylthiourea was assessed. Based on nominal concentrations, the 3-h EC50, EC10 and NOEC values without ATU were determined to be 103 mg test item L-1 (40.33 mg Pt L-1), 6.0 mg test item L-1 (2.35 mg Pt L-1), and 3.2 mg test item L-1 (1.26 mg Pt L-1), respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 83 mg test item L-1 (32.49 mg Pt L-1), 8.2 mg test item L-1 (3.21 mg Pt L-1), and 10 mg test item L-1 (3.92 mg Pt L-1), respectively.

Executive summary:

The inhibition of the respiration of activated sludge when exposed to the test item hexachloroplatinic acid was investigated. The study is reliable without restrictions, being GLP-compliant and having followed the standard test guidelines (OECD 209 and EU-Method C.11).

Two test series using the same concentrations were carried out, with and without the nitrification inhibitor N- allylthiourea (ATU) to discern between inhibition of nitrificators and inhibition of the total population. Because significant inhibition was observed, two additional experiments with and without ATU, respectively, were performed under the same test conditions. A positive control was used, and all validity criteria were met.

Based on nominal concentrations, the 3-h EC50, EC10 and NOEC values without ATU were determined to be 103 mg test item L^-1(40.33 mg Pt L^-1), 6.0 mg test item L^-1(2.35 mg Pt L^-1), and 3.2 mg test item L^-1(1.26 mg Pt L^-1), respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 83 mg test item L^-1(32.49 mg Pt L^-1), 8.2 mg test item L^-1(3.21 mg Pt L^-1), and 10 mg test item L^-1(3.92 mg Pt L^-1), respectively.

Description of key information

The 3-h EC50, EC10 and NOEC without ATU (nitrification inhibitor) were determined to be 188.65 mg Pt L^-1, 20.58 mg Pt L^-1 and 10.29 mg Pt L^-1, respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 240.1 mg Pt L^-1, 29.16 mg Pt L^-1 and 22.3 mg Pt L^-1, respectively. The key study driving the PNEC STP for platinum(IV) substances is an activated sludge respiration inhibition study for hexachloroplatinic acid. The 3-h EC50, EC10 and NOEC values without ATU were determined to be 40.33 mg Pt L^-1, 2.35 mg Pt L^-1 and 1.26 mg Pt L^-1, respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 32.49 mg Pt L^-1, 3.21 mg Pt L^-1 and 3.92 mg Pt L^-1, respectively.

Additional information

An activated sludge respiration inhibition test was conducted with dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol following OECD guideline 209 and EU Method C.11 (Simon 2016). Two test series using the same concentrations were carried out, with and without the nitrification inhibitor N- allylthiourea (ATU) to discern between inhibition of nitrificators and inhibition of the total population. Because significant inhibition was observed, two additional experiments with and without ATU, respectively, were performed under the same test conditions. A positive control was used, and all validity criteria were met. Based on nominal concentrations, the 3-h EC50, EC10 and NOEC without ATU were determined to be 1100 mg test item L^-1 (188.65 mg Pt L^-1), 120 mg test item L^-1 (20.58 mg Pt L^-1), and 60 mg test item L^-1 (10.29 mg Pt L^-1), respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 1400 mg test item L^-1 (240.1 mg Pt L^-1), 170 mg test item L^-1 (29.16 mg Pt L^-1), and 130 mg test item L^-1 (22.3 mg Pt L^-1), respectively.

 

For platinum(IV) substances there is some evidence that substances containing a chloro ligand are more toxic than those without a chloro ligand. For this reason, for substances without a chloro ligand, read across for REACH endpoints and for classification purposes is only conducted with other substances that do not contain a chloro ligand. However, for risk assessment purposes measured platinum concentrations in the environment only allow assessment of total platinum concentrations and do not differentiate between the form of the platinum in the environment. The PNEC for platinum(IV) substances is therefore based on pooled ecotoxicity data for all platinum(IV) substances, regardless of the ligands, and is based on the most toxic substance. For platinum(IV) substances the most toxic substance is hexachloroplatinic acid and the PNECs for platinum(IV) substances are therefore based on data for this substance. The key study driving the PNEC STP for platinum(IV) substances is an activated sludge respiration inhibition study for hexachloroplatinic acid.

 

An activated sludge respiration inhibition test was conducted with hexachloroplatinic acid following OECD guideline 209 and EU Method C.11 (Muckle 2015). Two test series were carried out, with and without the nitrification inhibitor N- allylthiourea (ATU) to discern between inhibition of nitrificators and inhibition of the total population. Based on a range-finding test where significant inhibition was observed, two additional tests were performed, where 5 concentrations ranging from 1 to 320 mg test item L^-1 were tested with and without ATU, respectively. A positive control was used, and all validity criteria were met. Based on nominal concentrations, the 3-h EC50, EC10 and NOEC values without ATU were determined to be 103 mg test item L^-1 (40.33 mg Pt L^-1), 6.0 mg test item L^-1 (2.35 mg Pt L^-1), and 3.2 mg test item L^-1 (1.26 mg Pt L^-1), respectively. The 3-h EC50, EC10 and NOEC values with ATU were determined to be 83 mg test item L^-1 (32.49 mg Pt L^-1), 8.2 mg test item L^-1 (3.21 mg Pt L^-1), and 10 mg test item L^-1 (3.92 mg Pt L^-1), respectively.