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Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10 August 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP, carried out according to recognised guideline
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.11 (Biodegradation: Activated Sludge Respiration Inhibition Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 850.6800 (Modified Activated Sludge, Respiration Inhibition Test for Sparingly Soluble Chemicals)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material:
Not applicable
Analytical monitoring:
no
Details on sampling:
Not applicable
Vehicle:
no
Details on test solutions:
Defined amounts of test item were directly weighed into the designated test flasks to reach the planned nominal concentrations. The nominal test item concentrations were prepared by mechanical dispersion using shortly (5 min) ultrasonic bath. These test solutions were freshly prepared at the beginning of the experiment, in the testing laboratory.
Test organisms (species):
activated sludge, domestic
Details on inoculum:
The activated sludge was supplied from the sewage plant for domestic sewage in Veszprém, Hungary.
The activated sludge used for this study was washed and centrifuged and the supernatant liquid phase was decanted. The solid material was re-suspended in isotonic saline solution and again centrifuged. This procedure was repeated twice.
An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to dry weight determined.
Based on this ratio, calculated amounts of wet sludge were suspended in isotonic saline solution to yield a concentration equivalent to 4 g per litre (on dry weight basis). The pH of the activated sludge inoculum was determined to be pH 7.20. The activated sludge was used directly after conditioning.
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
3 h
Post exposure observation period:
None
Hardness:
Not reported
Test temperature:
19.5 – 20.6 deg C (during the incubation) and
19.8 – 20.9 deg C (during oxygen measurement)
pH:
Not reported
Dissolved oxygen:
The concentration of dissolved oxygen did not drop below 2.5 mg O2/L during the incubation period, and just before the measurements of the respiration rates the oxygen concentrations were at least 7.0 mg O2/L.
Salinity:
Not applicable.
Nominal and measured concentrations:
The test concentrations (10, 31, 100, 313 and 1000 mg/L) were chosen to permit the determination of the EC50. Concentrations in excess of nominal 1000 mg test item/L were not tested.
Details on test conditions:
Surrounding type: Climate chamber (during the incubation) and controlled environment room (during the formulation and oxygen measuring)
Temperature: 19.5 – 20.6 deg C (during the incubation) and 19.8 – 20.9 deg C (during oxygen measurement)
Aeration: With compressed air (1 litre per minute)
Recording: Test conditions were measured with suitable instruments and documented in the raw data.

CONTROLS
Untreated Control (C1 and C2): Two controls (deionised water, synthetic sewage and inoculum, but without addition of the test item) were tested in parallel.
Reference Control (R1 – R3): In parallel to the study with the test item, the reference item 3,5-Dichlorophenol was tested (the nominal test concentrations of 5, 16 and 32 mg/L) under otherwise identical test conditions. A stock solution of 3,5-Dichlorophenol was prepared according to the OECD Guideline No. 209: 0.25 g of 3,5-Dichlorophenol was dissolved in 5 mL 1 mol/L NaOH and diluted to about 15 mL with deionised water. Excess of NaOH was neutralised with approximately 4 mL of 0.5 mol/L H2SO4 to the point of incipient precipitation. Thereafter, the mixture was made up to with deionised water. The final pH was measured to be 7.62 and the final concentration amounted 500 mg/L.
 
TEST UNITS
Type and size: Erlenmeyer bottles of approximately 350 mL volume, and BOD bottles with special neck of 300 mL volume.Identification: Each test flask was uniquely identified with at least study code, treatment and replicate codes (in case of controls).
 
PERFORMANCE OF THE TEST
Preparations of the test flasks: One test solution with a final volume of 330 mL was tested per treatment in a glass flask. 10.56 mL synthetic sewage and an adequate amount of the test item or an adequate volume of the stock solution of the reference item was filled up with deionised water to 198 mL before the start of the test. At the start of the test 132 mL activated sludge inoculum with a sludge concentration of 4 g/L (dry weight) was added, first to first control (C1), then in time intervals of 15 minutes (an arbitrary but convenient interval) to the test solutions of the reference item and the test item and finally to a second control (C2). Time interval between the third reference item flask and the first test item solution flask was more than 15 minutes.

Measurement of Respiration Rate
  
For the measurement of the respiration rate a well-mixed sample of each treatment was poured into a BOD flask after exactly 3 hours incubation time, and was not further aerated. The oxygen concentration was measured with a stirring O2electrode and was recorded for about ten minutes. The oxygen consumption (in mg O2L-1minute-1) was determinedfrom the most linear part of the respiration curve.
 
Measurement of pH, Dissolved Oxygen and Water Temperature
  
The pH and the oxygen concentrations were determined at the start and at the end of the incubation period in all treatments. The temperature was measured in the climate chamber with a min/max thermometer during the incubation period. The water temperature was recorded during the oxygen measurement in all BOD bottles.


Reference substance (positive control):
yes
Remarks:
3,5-Dichlorophenol
Key result
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
Key result
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:
In comparison to the inoculum controls the respiration rate of the activated sludge was inhibited between 4.2 % and 12.5 % in the examined nominal test concentration range (10 - 1000 mg/L). Concentrations exceeding 1000 mg/L nominal were not tested.
The influence of Kronitex TXP on the respiration rate of activated sludge is shown in Table 1 and in Figure 1.
Based on measured inhibition rates it can be stated that the 3-hour EC20, EC50 and EC80 were higher than 1000 mg/L.
The NOEC was determined to be 1000 mg/L.
The NOEC for respiration rates was not based on the results of a statistical analysis, but it is biologically justified. The respiration rates were inhibited and influenced dose dependently in the whole concentration range, and the observed slight inhibition (12.5 %) at the concentration level of 1000 mg/L was evaluated as reflecting the biological variability in the test. The calculated respiration rate, 0.420 (at 1000 mg/L) was not in the historical control data range (0.524 +/- 0.070), but the deviation from the control was within +/- 15 %, it can be considered as a biological variability of the test system.
Results with reference substance (positive control):
The following nominal concentrations of the positive reference control 3,5-Dichlorophenol were tested on the same activated sludge and under identical conditions as the test item: 5, 16 and 32 mg/L. In comparison to the controls the respiration rate of the activated sludge was inhibited by 35.4 % at the lowest nominal concentration of 5 mg/L. At the nominal concentrations of 16 and 32 mg/L, the respiration rate was inhibited by 60.4 % and 81.3 %, respectively. The 3-hour EC50 of 3,5-Dichlorophenol was calculated to be 9.46 mg/L with 95 % confidence limits of 7.40 to 12.11 mg
Reported statistics and error estimates:
See above.

Table 1.     Influence of test item on oxygen consumption of activated sludge

Flask
No.

ID

Test group

Concentration
(mg/L)

Oxygen consumption
(mg O2/L/min)

Inhibition
(%)

pH-values

Oxygen concentration
(mg O2/L)

start *

end *

start *

end *

1

C1

Control

0.490

7.26

7.31

8.1

8.0

15

C2

Control

0.470

7.29

7.36

8.4

7.9

 

Mean

0.480

 

Deviation (%)

4.2

10

T1

Test item

10

0.460

4.2

7.20

7.60

8.3

7.3

11

T2

Test item

31

0.450

6.3

7.25

7.64

8.3

8.4

12

T3

Test item

100

0.450

6.3

7.20

7.65

8.2

7.4

13

T4

Test item

313

0.430

10.4

7.15

7.56

8.3

7.7

14

T5

Test item

1000

0.420

12.5

7.52

7.65

8.4

7.0

Table 2.         Influence of test item on oxygen consumption of activated sludge

 

Flask
No.

ID

Test group

Concentration
(mg/L)

Oxygen consumption
(mg O2/L/min)

Inhibition
(%)

pH-values

Oxygen concentration
(mg O2/L)

start *

end *

start *

end *

1

C1

Control

0.490

7.26

7.31

8.1

8.0

15

C2

Control

0.470

7.29

7.36

8.4

7.9

 

Mean

0.480

 

Deviation (%)

4.2

10

T1

Test item

10

0.460

4.2

7.20

7.60

8.3

7.3

11

T2

Test item

31

0.450

6.3

7.25

7.64

8.3

8.4

12

T3

Test item

100

0.450

6.3

7.20

7.65

8.2

7.4

13

T4

Test item

313

0.430

10.4

7.15

7.56

8.3

7.7

14

T5

Test item

1000

0.420

12.5

7.52

7.65

8.4

7.0

*     start and end of 3-hour aeration

ID:        Code of each group

 

VALIDITY CRITERIA -The respiration rates of the two controls did not differ by more than 15 % (4.2 %). -The 3-hour EC50 of the reference item 3,5-Dichlorophenol for the used activated sludge batch was determined to be in the range of 5 to 30 mg/L (9.46 mg/L). -The concentration of dissolved oxygen did not drop below 2.5 mg O2/L during the incubation period, and just before the measurements of the respiration rates the oxygen concentrations were at least 7.0 mg O2/L.

Validity criteria fulfilled:
yes
Conclusions:
A laboratory test was carried out with Kronitex TXP to evaluate the effect of the test item on microorganisms by measuring the respiration rate.
The test concentrations (10, 31, 100, 313 and 1000 mg/L) were chosen to permit the determination of the EC50.

In comparison to the inoculum controls the respiration rate of the activated sludge was inhibited between 4.2 % and 12.5 % in the examined nominal test concentration range (10 - 1000 mg/L). Concentrations exceeding 1000 mg/L nominal were not tested.

In parallel to the study with the test item, the reference item 3,5-Dichlorophenol was tested (the nominal test concentrations of 5, 16 and 32 mg/L) under otherwise identical test conditions.

The 3-hour EC50 of 3,5-Dichlorophenol was calculated to be 9.46 mg/L with 95 % confidence limits of 7.40 to 12.11 mg/L.


Based on measured inhibition rates it can be stated that the 3-hour EC20, EC50 and EC80 were higher than 1000 mg/L.
Executive summary:

In comparison to the inoculum controls the respiration rate of the activated sludge was inhibited between 4.2 % and 12.5 % in the examined nominal test concentration range (10 - 1000 mg/L). Concentrations exceeding 1000 mg/L nominal were not tested.

 

In parallel to the study with the test item, the reference item 3,5-Dichlorophenol was tested (the nominal test concentrations of 5, 16 and 32 mg/L) under otherwise identical test conditions.

 

The 3-hour EC50of 3,5-Dichlorophenol was calculated to be 9.46 mg/L with 95 % confidence limits of 7.40 to 12.11 mg/L.

Based on measured inhibition rates it can be stated that the 3-hour EC20, EC50and EC80were higher than 1000 mg/L.

The NOEC was determined to be 1000 mg/L.

The NOEC for respiration rates was not based on the results of a statistical analysis, but it is biologically justified. The respiration rates were inhibited and influenced dose dependently in the whole concentration range, and the observed slight inhibition (12.5 %) at the concentration level of 1000 mg/L was evaluated as reflecting the biological variability in the test. The calculated respiration rate, 0.420 (at 1000 mg/L) was not in the historical control data range (0.524±0.070), but the deviation from the control was within +/- 15 %, it can be considered as a biological variability of the test system.

Endpoint:
activated sludge respiration inhibition testing
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Please see the attached justification.
Reason / purpose:
read-across source
Key result
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
Remarks on result:
other: CAS 68937-41-7
Key result
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
Remarks on result:
other: CAS 68937-41-7
Validity criteria fulfilled:
yes
Conclusions:
The substance, CAS 25155-23-1; EC 246-677-8, is analogous to the substance to be read across to, in terms of basic form, and the degree of substitution of functional groups is not considered to effect the proposed read across for the endpoint of toxicity to microogranisms. The substance is considered to be not toxic to microorganisms for the defined endpoints on the basis of read across.
Executive summary:

The substance, CAS 25155-23-1; EC 246-677-8, is analogous to the substance to be read across to, in terms of basic form, and the degree of substitution of functional groups is not considered to effect the proposed read across for the endpoint of toxicity to microogranisms. The substance is considered to be not toxic to microorganisms for the defined endpoints on the basis of read across.

Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10 August 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant, carried out according to recognised guidelines
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
yes
Remarks:
see below
Qualifier:
according to
Guideline:
EU Method C.11 (Biodegradation: Activated Sludge Respiration Inhibition Test)
Deviations:
yes
Remarks:
see below
Qualifier:
according to
Guideline:
EPA OPPTS 850.6800 (Modified Activated Sludge, Respiration Inhibition Test for Sparingly Soluble Chemicals)
Deviations:
yes
Remarks:
see below
Principles of method if other than guideline:
DEVIATION FROM THE STUDY PLAN

Concerning: Preparations of the test flasks

According to the Study Plan: At the start of the test 132 mL activated sludge inoculum with a sludge concentration of 2-4 g/L (dry weight) will be added, first to first control (C1), then in time intervals of approximately 15 minutes (an arbitrary but convenient interval) to the test solutions of the reference item and the test item and finally to a second control (C2).

Deviation: At the start of the test 132 mL activated sludge inoculum with a sludge concentration of 4 g/L (dry weight) was added, first to first control (C1), then in time intervals of 15 minutes (an arbitrary but convenient interval) to the test solutions of the reference item and the test item and finally to a second control (C2). Time interval between the last test solution flask and the second control was more than 15 minutes.

Reason for this change: Technical
Presumed Effect on the Study: None
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material:
Not applicable
Analytical monitoring:
no
Details on sampling:
Not applicable
Vehicle:
no
Details on test solutions:
The test concentrations (10, 31, 100, 313 and 1000 mg/L) were chosen to permit the determination of the EC50.

Defined amounts of test item were directly weighed into the designated test flasks to reach the planned nominal concentrations. The nominal test item concentrations were prepared by mechanical dispersion using shortly (5 min) ultrasonic bath. These test solutions were freshly prepared at the beginning of the experiment, in the testing laboratory.
Untreated Control (C1 and C2): Two controls (deionised water, synthetic sewage and inoculum, but without addition of the test item) were tested in parallel.
Reference Control (R1 – R3): In parallel to the study with the test item, the reference item 3,5-Dichlorophenol was tested (the nominal test concentrations of 5, 16 and 32 mg/L) under otherwise identical test conditions. A stock solution of 3,5-Dichlorophenol was prepared according to the OECD Guideline No. 209: 0.25 g of 3,5-Dichlorophenol was dissolved in 5 mL 1 mol/L NaOH and diluted to about 15 mL with deionised water. Excess of NaOH was neutralised with approximately 4 mL of 0.5 mol/L H2SO4 to the point of incipient precipitation. Thereafter, the mixture was made up to 0.5 litre with deionised water. The final pH was measured to be 7.62 and the final concentration amounted 500 mg/L.

Synthetic Sewage Feed (ratio of composition of culture media referring to 1000 mL)

Peptone 16.0g
Meat extract 11.0g
Urea 3.0g
NaCl 0.7g
CaCl2 x 2H2O 0.4g
MgSO4 x 7H2O 0.2g
K2HPO4 2.8g
Deionised water add 1000.0mL

Test organisms (species):
activated sludge, domestic
Details on inoculum:
Test organisms comprised activated sludge, microorganisms from a domestic waste water treatment plant.
The activated sludge was supplied from the sewage plant for domestic sewage in Veszprém, Hungary.
The activated sludge used for this study was washed and centrifuged and the supernatant liquid phase was decanted. The solid material was re-suspended in isotonic saline solution and again centrifuged. This procedure was repeated twice. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to dry weight determined. Based on this ratio, calculated amounts of wet sludge were suspended in isotonic saline solution to yield a concentration equivalent to 4 g per litre (on dry weight basis). The pH of the activated sludge inoculum was determined to be pH 7.20. The activated sludge was used directly after conditioning.
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
3 h
Post exposure observation period:
None
Hardness:
Not measured
Test temperature:
Room temperature (20-25 deg C)
pH:
7.21 - 7.87
Dissolved oxygen:
7.0 - 8.5
Salinity:
Not applicable.
Conductivity:
Not specified
Nominal and measured concentrations:
10, 31, 100, 313, 1000 mg/l
Details on test conditions:
Surrounding type: Climate chamber (during the incubation) and controlled environment room (during the formulation and oxygen measuring)
Temperature: 19.5 – 20.6 deg C (during the incubation) and 20.0 – 20.9 deg cC (during oxygen measurement)
Aeration: With compressed air (1 litre per minute)
Recording: Test conditions were measured with suitable instruments and documented in the raw data.

Reference substance (positive control):
yes
Remarks:
3,5-Dichlorophenol (the nominal test concentrations of 5, 16 and 32 mg/L) under otherwise identical test conditions.
Key result
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
Key result
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:
In comparison to the inoculum controls the respiration rate of the activated sludge was not inhibited in the concentration of 10 mg Reofos 65/L. In comparison to the inoculum controls the respiration rate of the activated sludge was inhibited between 2.1 % and 4.2 % in the examined nominal test concentration range (31 - 1000 mg/L). Concentrations exceeding 1000 mg/L nominal were not tested.
Based on measured inhibition rates it can be stated that the 3-hour EC20, EC50and EC80were higher than 1000 mg/L.
The NOEC was determined to be 1000 mg/L. The NOEC for respiration rates was not based on the results of a statistical analysis, but it is biologically justified. The respiration rates were inhibited and influenced dose dependently in the whole concentration range, and the observed slight inhibition (4.2 %) at the concentration level of 1000 mg/L was evaluated as reflecting the biological variability in the test. The calculated respiration rate, 0.460 (at 1000 mg/L) was not in the historical control data range (0.524 ±0.070), but the deviation from the control was within +/- 15 %, it can be considered as a biological variability of the test system.
Results with reference substance (positive control):
The following nominal concentrations of the positive reference control 3,5-Dichlorophenol were tested on the same activated sludge and under identical conditions as the test item: 5, 16 and 32 mg/L. In comparison to the controls the respiration rate of the activated sludge was inhibited by 35.4 % at the lowest nominal concentration of 5 mg/L. At the nominal concentrations of 16 and 32 mg/L, the respiration rate was inhibited by 60.4 % and 81.3 %, respectively.
The 3-hour EC50 of 3,5-Dichlorophenol was calculated to be 9.46 mg/L with 95 % confidence limits of 7.40 to 12.11 mg/L.

Table 2.     Influence of test item on oxygen consumption of activated sludge

Flask
No.

ID

Test group

Concentration
(mg/L)

Oxygen consumption
(mg O2/L/min)

Inhibition
(%)

pH-values

Oxygen concentration
(mg O2/L)

start *

end *

start *

end *

1

C1

Control

0.490

7.26

7.31

8.1

8.0

15

C2

Control

0.470

7.29

7.36

8.4

7.9

 

Mean

0.480

 

Deviation (%)

4.2

5

T1

Test item

10

0.480

0.0

7.28

7.82

8.4

7.3

6

T2

Test item

31

0.470

2.1

7.26

7.87

8.5

7.6

7

T3

Test item

100

0.470

2.1

7.27

7.80

8.4

7.7

8

T4

Test item

313

0.470

2.1

7.32

7.80

8.5

7.0

9

T5

Test item

1000

0.460

4.2

7.21

7.79

8.4

7.0

Validity Criteria 

-The respiration rates of the two controls did not differ by more than 15 % (4.2 %).

-The 3-hour EC50of the reference item 3,5-Dichlorophenol for the used activated sludge batch was determined to be in the range of 5 to 30 mg/L (9.46 mg/L).

-The concentration of dissolved oxygen did not drop below 2.5 mg O2/L during the incubation period, and just before the measurements of the respiration rates the oxygen concentrations were at least 7.0 mg O2/L.

Validity criteria fulfilled:
yes
Conclusions:
This test was conducted on a sample of the material where the content of Triphenyl phosphate is present at > 5%

Based on measured inhibition rates it can be stated that the 3-hour EC20, EC50and EC80were higher than 1000 mg/L. The substance does not inhibit the respiration rate of sewage sludge microorganisms, and is not considered to be harmful to organisms of this nature. No classification is applicable.
Executive summary:

This test was conducted on a sample of the material where the content of Triphenyl phosphate is present at > 5%

A laboratory test was carried out with Reofos 65 to evaluate the effect of the test item on microorganisms by measuring the respiration rate. The test concentrations (10, 31, 100, 313 and 1000 mg/L) were chosen to permit the determination of the EC50.

In comparison to the inoculum controls the respiration rate of the activated sludge was not inhibited in the concentration of 10 mg Reofos 65/L. In comparison to the inoculum controls the respiration rate of the activated sludge was inhibited between 2.1 % and 4.2 % in the examined nominal test concentration range (31 - 1000 mg/L). Concentrations exceeding 1000 mg/L nominal were not tested.

In parallel to the study with the test item, the reference item 3,5-Dichlorophenol was tested (the nominal test concentrations of 5, 16 and 32 mg/L) under otherwise identical test conditions. 

The 3-hour EC50of 3,5-Dichlorophenol was calculated to be 9.46 mg/L with 95 % confidence limits of 7.40 to 12.11 mg/L.

Based on measured inhibition rates it can be stated that the 3-hour EC20, EC50and EC80were higher than 1000 mg/L.

The NOEC was determined to be 1000 mg/L.

The NOEC for respiration rates was not based on the results of a statistical analysis, but it is biologically justified. The respiration rates were inhibited and influenced dose dependently in the whole concentration range, and the observed slight inhibition (4.2 %) at the concentration level of 1000 mg/L was evaluated as reflecting the biological variability in the test. The calculated respiration rate, 0.460 (at 1000 mg/L) was not in the historical control data range (0.524 ± 0.070), but the deviation from the control was within +/- 15 %, it can be considered as a biological variability of the test system.

Description of key information

Toxicity to microorganisms

Key value for chemical safety assessment

EC50 for microorganisms:
1 000 mg/L
EC10 or NOEC for microorganisms:
1 000 mg/L

Additional information

Experimental study using the test substance.

In comparison to the inoculum controls the respiration rate of the activated sludge was inhibited between 4.2 % and 12.5 % in the examined nominal test concentration range (10 - 1000 mg/L). Concentrations exceeding 1000 mg/L nominal were not tested.

 

In parallel to the study with the test item, the reference item 3,5-Dichlorophenol was tested (the nominal test concentrations of 5, 16 and 32 mg/L) under otherwise identical test conditions.

 

The 3-hour EC50of 3,5-Dichlorophenol was calculated to be 9.46 mg/L with 95 % confidence limits of 7.40 to 12.11 mg/L.

 

Based on measured inhibition rates it can be stated that the 3-hour EC20, EC50and EC80were higher than 1000 mg/L.

 

The NOEC was determined to be 1000 mg/L.

 

The NOEC for respiration rates was not based on the results of a statistical analysis, but it is biologically justified. The respiration rates were inhibited and influenced dose dependently in the whole concentration range, and the observed slight inhibition (12.5 %) at the concentration level of 1000 mg/L was evaluated as reflecting the biological variability in the test. The calculated respiration rate, 0.420 (at 1000 mg/L) was not in the historical control data range (0.524±0.070), but the deviation from the control was within +/- 15 %, it can be considered as a biological variability of the test system.

Read-across information:

Two studies are presented for Activated Sludge Respiration Inhibition with the following results:

 

Where TPP content > 5%

NOEC (3 h): 1000 mg/L test mat. (nominal) based on: respiration rate

EC50 (3 h): > 1000 mg/L test mat. (nominal) based on: respiration rate

 

Where TPP content < 5%

EC50 (3 h): > 1000 ppm dissolved (nominal) based on: respiration rate

 

The substance is not considered to pose a hazard to STP microorganisms and is not dependent on triphenyl phosphate content.