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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

Study period:

The study was conducted between 11 March 2009 and 12 March 2009.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: guideline study on Gas-to-liquids (GTL) substance covering the carbon range from C18 to C50

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Method C.11 of Commission Regulation (EC) No. 440/2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: US EPA Draft Ecological Effects Test Guidelines OPPTS 850.6800

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Details on sampling:

- Concentrations:
10, 32, 100, 320 and 1000 mg/l

- Sampling method:
Observations were made on the test preparations throughout the test period. Observations of the test material vessels at 0 hours were made prior to addition of activated sewage sludge and synthetic sewage. The pH of the control, reference material and test material preparations were measured using a WTW pH/Oxi 340I pH and dissolved oxygen meter at 0 hours and prior to measurement of the oxygen consumption rate after 3 hours contact time.
- Sample storage conditions before analysis:
not specified in report

Vehicle:

no

Details on test solutions:

Test material preparation
For the purpose of the test, the test material was dispersed directly in water.
Amounts of test material (5, 16, 50, 160 and 500 mg) were each separately dispersed in approximately 250 ml of water and subjected to ultrasonication for approximately 15 minutes followed by prolonged mixing for 24 hours in order to maximise the dissolved test material concentration. All test vessels were shielded from the light during mixing. Synthetic sewage (16 ml), activated sewage sludge (200 ml) and water were added to a final volume of 500 ml to give the required concentrations of 10, 32, 100, 320 and 1000 mg/l.
The control group was maintained under identical conditions but not exposed to the test material.

Reference material preparation
For the purpose of the test a reference material, 3,5-dichlorophenol (Sigma-Aldrich Batch No. 04621CJ) was used. Two stock solutions of 50 and 160 mg/l were prepared by dissolving the reference material directly in water with the aid of ultrasonication for approximately 15 minutes. Aliquots (10 and 100 ml) of the 160 mg/l stock solution were removed and dispersed with activated sewage sludge, synthetic sewage and water to give the final concentrations of 3.2 and 32 mg/l. Similarly, a 100 ml aliquot of the 50 mg/l stock solution was used to prepare the 10 mg/l concentration. The volumetric flasks containing the reference material were inverted several times to ensure homogeneity of the solutions.

Synthetic Sewage
A synthetic sewage of the following composition, was added to each test vessel to act as a respiratory substrate:
16 g Peptone
11 g Meat extract
3 g Urea
0.7 g NaCl
0.4 g CaCl2.2H2O
0.2 g MgSO4.7H2O
2.8 g K2HPO4
dissolved in 1 litre of water with the aid of ultrasonication.



- Eluate:
At time "0" 16 ml of synthetic sewage was diluted to 300 ml with water and 200 ml of inoculum added in a 500 ml conical flask (first control). The mixture was aerated with clean, oil-free compressed air via narrow bore glass tubes at a rate of approximately 0.5 – 1 litre per minute. Thereafter, at 15 minute intervals the procedure was repeated with appropriate amounts of the reference material being added. The test material vessels were prepared as described above. Finally a second control was prepared.
As each vessel reached 3 hours contact time an aliquot was removed from the conical flask and poured into the measuring vessel (250 ml darkened glass Biological Oxygen Demand (BOD) bottle) and the rate of respiration measured using a Yellow Springs dissolved oxygen meter fitted with a BOD probe. The contents of the measuring vessel were stirred constantly by magnetic stirrer. The rate of respiration for each flask was measured over the linear portion of the oxygen consumption trace (where possible between approximately 6.5 mg O2/l and 2.5 mg O2/l). In the case of a rapid oxygen consumption, measurements may have been outside this range but the oxygen consumption was always within the linear portion of the respiration curve. In the case of low oxygen consumption, the rate was determined over an approximate 10 minute period.
The test was conducted under normal laboratory lighting in a temperature controlled room at 21±1DegC.

- Differential loading:
Not applicable.

- Controls:
The control group was maintained under identical conditions but not exposed to the test material.

- Chemical name of vehicle (organic solvent, emulsifier or dispersant):
Not applicable.

- Concentration of vehicle in test medium (stock solution and final test solution(s) including control(s)):
Not applicable.

- Evidence of undissolved material (e.g. precipitate, surface film, etc):
Observations made at 0 hours (see Table 3 attached in overall remarks and attachments) prior to the addition of activated sewage sludge and synthetic sewage showed that all test concentrations an oily layer of test material was visible on the surface of a clear colourless water column.
This was considered to be due to the insoluble nature of the test material in the test media. However the test material was dispersed with the aid of ultrasonication in the test diluent for approximately 15 minutes prior to prolonged stirring for 24 hours. Each vessel was also aerated with compressed air at a rate of approximately 0.5-1 litre per minute to ensure that there was maximum contact between the test material and activated sewage sludge in the test medium.
Observations made after 30 minutes and 3 hours contact time showed that all test concentrations in a dark brown dispersion an oily layer of test material was visible.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

Test Species
A mixed population of activated sewage sludge micro-organisms was obtained on 12 March 2009 from the aeration stage of the Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, UK which treats predominantly domestic sewage.

- Laboratory culture:
The activated sewage sludge sample was maintained on continuous aeration in the laboratory at a temperature of approximately 21ºC and was used on the day of collection. The pH of the sample was 7.9 measured using a WTW pH/Oxi 340I pH and dissolved oxygen meter. Determination of the suspended solids level of the activated sewage sludge was carried out by filtering a sample (100 ml) of the activated sewage sludge by suction through a pre-weighed GF/A filter paper using a Buchner funnel which was then rinsed 3 times with 10 ml of deionised reverse osmosis water and filtration continued for 3 minutes. The filter paper was then dried in an oven at approximately 105ºC for at least 1 hour and allowed to cool before weighing. This process was repeated until a constant weight was attained. The suspended solids concentration was equal to 3.7 g/l prior to use.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Post exposure observation period:

Not applicable.

Hardness:

The test water used for the test was laboratory tap water dechlorinated by passage through an activated carbon filter (Purite Series 500) and partly softened (Elga Nimbus 1248D Duplex water softener) giving water with a total hardness of approximately 140 mg/l as CaCO3. After dechlorination and softening the water was then passed through a series of computer controlled plate heat exchangers to achieve the required temperature. Typical water quality characteristics for the tap water as supplied, prior to dechlorination and softening, are given in Appendix 1 (attached in overall remarks and attachments).

Test temperature:

The test was conducted under normal laboratory lighting in a temperature controlled room at 21±1DegC.

pH:

The pH values of the test preparations at the start and end of the exposure period are given in Table 2 (see any other information on materials and results, including tables section)

Dissolved oxygen:

In some instances, the initial and final dissolved oxygen concentrations were below those recommended in the test guidelines (6.5 mg O2/l and 2.5 mg O2/l respectively). This was considered to have had no adverse effect on the results of the study given that in all cases the oxygen consumption rate was determined over the linear portion of the oxygen consumption trace.

Salinity:

Not applicable.

Nominal and measured concentrations:

10, 32, 100, 320 and 1000 mg/l

Details on test conditions:

OTHER TEST CONDITIONS
- Adjustment of pH: The pH values of the test preparations at the start and end of the exposure period are given in Table 2 (see any other information on materials and results, including tables section).

- Photoperiod: 3 hours.

- Light intensity: Normal laboratory lighting.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
In order to calculate the inhibitory effect of the test and reference materials the respiration rate was expressed as a percentage of the two control respiration rates.

% inhibition = [ 1 – 2 RS ] x 100
RC1 + RC2
where
RS = oxygen consumption rate for test or reference sample
RC1 + RC2 = oxygen consumption rates for controls 1 and 2

The percentage inhibition values were plotted against concentration for the reference material only, a line fitted using the Xlfit software package (IDBS) and the EC15, EC20, EC50 and EC80 values determined from the equation for the fitted line.
The EC15, EC20, EC50 and EC80 values for the test material were determined by inspection of the inhibition of respiration rate data.
95% confidence limits were calculated for the reference material EC50 value using the method of Litchfield and Wilcoxon (Litchfield and Wilcoxon 1949).
The No Observed Effect Concentration (NOEC) was taken as being the EC15 value.
The results of the study are considered valid if (i) the two control respiration rates are within 15% of each other and (ii) the EC50 (3-Hour contact time) for 3,5-dichlorophenol lies within the range 5 to 30 mg/l.


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

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: not specified

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: not specified

Details on results:

Oxygen consumption rates and percentage inhibition values for the control, test and reference materials are given in Table 1 (see any other information on materials and results, including tables section) . The pH values of the test preparations at the start and end of the exposure period are given in Table 2 (see any other information on materials and results, including tables section), and observations made on the test preparations throughout the study are given in Table 3 (see any other information on materials and results, including tables section)
Percentage inhibition is plotted against concentration for the reference material, 3,5 dichlorophenol only (Figure 1 attached in overall remakrs and attachments section).
The following results were derived:

ECx (3 Hours) (mg/l) 95% Confidence Limits (mg/l)
EC20 >1000 -
EC50 >1000 -
EC80 >1000 -
NOEC 1000 -

It was considered unnecessary and unrealistic to test at concentrations in excess of 1000 mg/l.

3,5-dichlorophenol:
ECx (3 Hours) (mg/l) 95% Confidence Limits (mg/l)
EC20 2.6 -
EC50 7.9 6.1 - 10
EC80 25 -
NOEC 2.1
Variation in respiration rates of controls 1 and 2 after 3 hours contact time was ± 5%.
The validation criteria for the control respiration rates and reference material EC50 values were therefore satisfied.

Results with reference substance (positive control):

- Results with reference substance valid?
Yes.

- Relevant effect levels:
The reference material gave a 3-Hour EC50 value of 7.9 mg/l, 95% confidence limits 6.1 - 10 mg/l.

- Other:
None.

Reported statistics and error estimates:

None.

REFERENCES

Litchfield, J T and Wilcoxon, F (1949) A Simplified Method of Evaluating Dose-Effect Experints. J Pharmacol Exp Ther 96, 99-113.

Xlfit, ID Business Solutions Ltd.

Validity criteria fulfilled:

yes

Conclusions:

The effect of the test material on the respiration of activated sewage sludge micro-organisms gave a 3-Hour EC50 of greater than 1000 mg/l. The No Observed Effect Concentration (NOEC) after 3 hours exposure was 1000 mg/l.

Executive summary:

Introduction. A study was perford to assess the effect of the test material 'Distillates (Fischer-Tropsch), heavy, C18-50 - branched, cyclic and linear’ on the respiration of activated sewage sludge. Thethod followed that described in the OECD Guidelines for Testing of Chemicals (1984) No 209 "Activated Sludge, Respiration Inhibition Test", Method C.11 of Commission Regulation (EC) No. 440/2008 and US EPA Draft Ecological Effects Test Guidelines OPPTS 850.6800.

Methods. Activated sewage sludge was exposed to an aqueous dispersion of the test material at concentrations of 10, 32, 100, 320 and 1000 mg/l for a period of 3 hours at a temperature of approximately 21°C with the addition of a synthetic sewage as a respiratory substrate.

The rate of respiration was determined after 3 hours contact tiand compared to data for the control and a reference material, 3,5-dichlorophenol.

Results. The effect of the test material on the respiration of activated sewage sludge gave a 3‑Hour EC₅₀of greater than 1000 mg/l. The No Observed Effect Concentration (NOEC) after 3 hours exposure was1000mg/l.

It was considered unnecessary and unrealistic to test at concentrations in excess of 1000 mg/l.

The reference material gave a 3-Hour EC₅₀value of 7.9 mg/l, 95% confidence limits 6.1 – 10 mg/l.

Conclusion.

The effect of the test material on the respiration of activated sewage sludge gave a 3‑Hour EC₅₀of greater than 1000 mg/l. The No Observed Effect Concentration (NOEC) after 3 hours exposure was1000mg/l.

It was considered unnecessary and unrealistic to test at concentrations in excess of 1000 mg/l.

The reference material gave a 3-Hour EC₅₀value of 7.9 mg/l, 95% confidence limits 6.1 – 10 mg/l.

Reference 2

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The study was conducted between 23 March 2009 and 24 March 2009.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: guideline study on Gas-to-liquids (GTL) substance covering the carbon range from C8 to C26

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

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.6800 (Modified Activated Sludge, Respiration Inhibition Test for Sparingly Soluble Chemicals)

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Not Applicable

Analytical monitoring:

no

Details on sampling:

Not appliciable

Vehicle:

no

Details on test solutions:

Test material preparation
For the purpose of the test, the test material was dispersed directly in water.

Amounts of test material (5, 16, 50, 160 and 500 mg) were each separately dispersed in approximately 250 ml of water and subjected to ultrasonication for approximately 15 minutes followed by prolonged mixing for 24 hours in order to maximise the dissolved test material concentration. All test vessels were shielded from the light during mixing. Synthetic sewage (16 ml), activated sewage sludge (200 ml) and water were added to a final volume of 500 ml to give the required concentrations of 10, 32, 100, 320 and 1000 mg/l.

The control group was maintained under identical conditions but not exposed to the test material.

Analysis of the concentration, homogeneity and stability of the test material in the test preparations was not appropriate to the Test Guideline.

Reference material preparation
For the purpose of the test a reference material, 3,5-dichlorophenol (Sigma-Aldrich Batch No. 04621CJ) was used. Two stock solutions of 50 and 160 mg/l were prepared by dissolving the reference material directly in water with the aid of ultrasonication for approximately 20 minutes. Aliquots (10 and 100 ml) of the 160 mg/l stock solution were removed and dispersed with activated sewage sludge, synthetic sewage and water to give the final concentrations of 3.2 and 32 mg/l. Similarly, a 100 ml aliquot of the 50 mg/l stock solution was used to prepare the 10 mg/l concentration. The volumetric flasks containing the reference material were inverted several times to ensure homogeneity of the solutions.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

Test Species
A mixed population of activated sewage sludge micro-organisms was obtained on 24 March 2009 from the aeration stage of the Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, UK which treats predominantly domestic sewage.
Preparation of inoculum
The activated sewage sludge sample was maintained on continuous aeration in the laboratory at a temperature of approximately 21ºC and was used on the day of collection. The pH of the sample was 7.4 measured using a WTW pH/Oxi 340I pH and dissolved oxygen meter. Determination of the suspended solids level of the activated sewage sludge was carried out by filtering a sample (100 ml) of the activated sewage sludge by suction through a pre-weighed GF/A filter paper using a Buchner funnel which was then rinsed 3 times with 10 ml of deionised reverse osmosis water and filtration continued for 3 minutes. The filter paper was then dried in an oven at approximately 105ºC for at least 1 hour and allowed to cool before weighing. This process was repeated until a constant weight was attained. The suspended solids concentration was equal to 3.9 g/l prior to use.


Synthetic Sewage
A synthetic sewage of the following composition, was added to each test vessel to act as a respiratory substrate:
16 g Peptone
11 g Meat extract
3 g Urea
0.7 g NaCl
0.4 g CaCl2.2H2O
0.2 g MgSO4.7H2O
2.8 g K2HPO4
dissolved in 1 litre of water with the aid of ultrasonication.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Post exposure observation period:

Not applicable.

Hardness:

The test water used for the test was laboratory tap water dechlorinated by passage through an activated carbon filter (Purite Series 500) and partly softened (Elga Nimbus 1248D Duplex water softener) giving water with a total hardness of approximately 140 mg/l as CaCO3. After dechlorination and softening the water was then passed through a series of computer controlled plate heat exchangers to achieve the required temperature. Typical water quality characteristics for the tap water as supplied, prior to dechlorination and softening, are given in Appendix 1 (in attached section).

Test temperature:

The test was conducted under normal laboratory lighting in a temperature controlled room at 21±1 DegC.

pH:

The pH of the control, reference material and test material preparations were measured using a WTW pH/Oxi 340I pH and dissolved oxygen meter at 0 hours and prior to measurement of the oxygen consumption rate after 3 hours contact time. (see Table 2 in section any other information onmaterials and methods.

Dissolved oxygen:

In some instances, the initial and final dissolved oxygen concentrations were below those recommended in the test guidelines (6.5 mg O2/l and 2.5 mg O2/l respectively). This was considered to have had no adverse effect on the results of the study given that in all cases the oxygen consumption rate was determined over the linear portion of the oxygen consumption trace.

Salinity:

Not applicable

Nominal and measured concentrations:

1000 mg/l.

Details on test conditions:

Preparation of test system
At time "0" 16 ml of synthetic sewage was diluted to 300 ml with water and 200 ml of inoculum added in a 500 ml conical flask (first control). The mixture was aerated with clean, oil-free compressed air via narrow bore glass tubes at a rate of approximately
0.5 – 1 litre per minute. Thereafter, at 15 minute intervals the procedure was repeated with appropriate amounts of the reference material being added.

Finally a second control was prepared.

As each vessel reached 3 hours contact time an aliquot was removed from the conical flask and poured into the measuring vessel (250 ml darkened glass Biological Oxygen Demand (BOD) bottle) and the rate of respiration measured using a Yellow Springs dissolved oxygen meter fitted with a BOD probe. The contents of the measuring vessel were stirred constantly by magnetic stirrer. The rate of respiration for each flask was measured over the linear portion of the oxygen consumption trace (where possible between approximately 6.5 mg O2/l and 2.5 mg O2/l). In the case of a rapid oxygen consumption, measurements may have been outside this range but the oxygen consumption was always within the linear portion of the respiration curve. In the case of low oxygen consumption, the rate was determined over an approximate 10 minute period.

Observations were made on the test preparations throughout the test period. Observations of the test material vessels at 0 hours were made prior to addition of activated sewage sludge and synthetic sewage.

Evaluation of Data
In order to calculate the inhibitory effect of the test and reference materials the respiration rate was expressed as a percentage of the two control respiration rates.


% inhibition = ( 1 – (2 RS/RC1 + RC2) x 100

where
RS = oxygen consumption rate for test or reference sample
RC1 + RC2 = oxygen consumption rates for controls 1 and 2

The percentage inhibition values were plotted against concentration for the reference material only, a line fitted using the Xlfit software package .
(IDBS) and the EC15, EC20, EC50 and EC80 values determined from the equation for the fitted line.
The EC15, EC20, EC50 and EC80 values for the test material were determined by inspection of the inhibition of respiration rate data.
95% confidence limits were calculated for the reference material EC50 values using the method of Litchfield and Wilcoxon (Litchfield and Wilcoxon 1949).

The No Observed Effect Concentration (NOEC) was taken as being the EC15 value.

The results of the study are considered valid if (i) the two control respiration rates are within 15% of each other and (ii) the EC50 (3-Hour contact time) for 3,5-dichlorophenol lies within the range 5 to 30 mg/l.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

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: not stated

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: not stated

Details on results:

Oxygen consumption rates and percentage inhibition values for the control, test and reference materials are given in Table 1 (in section any other information on materials and methods). The pH values of the test preparations at the start and end of the exposure period are given in Table 2 (in section any other information on materials and methods), and observations made on the test preparations throughout the study are given in Table 3 (in section any other information on materials and methods).

Percentage inhibition is plotted against concentration for the reference material, 3,5 dichlorophenol only (Figure 1 in attached section).
The following results were derived:


The following results were derived:
Distillates (Fischer-Tropsch), C8-26-branched and linear:
ECx (3 Hours) (mg/l) 95% Confidence Limits (mg/l)
EC20 >1000 -
EC50 >1000 -
EC80 >1000 -
NOEC 1000 -

It was considered unnecessary and unrealistic to test at concentrations in excess of 1000 mg/l.

3,5-dichlorophenol:
ECx (3 Hours) (mg/l) 95% Confidence Limits (mg/l)
EC20 2.4 -
EC50 7.9 6.1 - 10
EC80 25 -
NOEC 2.0

Variation in respiration rates of controls 1 and 2 after 3 hours contact time was ± 4%.
The validation criteria for the control respiration rates and reference material EC50 values were therefore satisfied.
In some instances, the initial and final dissolved oxygen concentrations were below those recommended in the test guidelines (6.5 mg O2/l and 2.5 mg O2/l respectively). This was considered to have had no adverse effect on the results of the study given that in all cases the oxygen consumption rate was determined over the linear portion of the oxygen consumption trace.
Observations made at 0 hours (see Table 3 in section any other information on materials and methods) prior to the addition of activated sewage sludge and synthetic sewage showed that the test concentrations of 10 and 32 mg/l contained clear colourless water columns with an oily slick of test material on the surface. The test concentrations of 100, 320 and 1000 mg/l contained clear colourless water columns with oily globules of test material on the surface.

Observations made after 30 minutes and 3 hours contact time showed that the test concentrations of 10 and 32 mg/l contained dark brown dispersions with no undissolved test material visible. This was considered to be due to the small amounts of test material present (5 and 16 mg) being obscured by the dark brown activated sewage sludge. The test concentrations of 100, 320 and 1000 mg/l contained dark brown dispersions with an oily slick of test material on the surface.

The test material was dispersed with the aid of ultrasonication in the test diluent for approximately 15 minutes prior to prolonged mixing for 24 hours in order to maximise the dissolved test material concentration. After the addition of the synthetic sewage, activated sewage sludge and water, each vessel was aerated with compressed air at a rate of approximately 0.5-1 litre per minute to ensure that there was maximum contact between the test material and activated sewage sludge in the test medium.

Results with reference substance (positive control):

- Results with reference substance valid? Yes.
- Relevant effect levels: The reference material gave a 3-Hour EC50 value of 8.6 mg/l, 95% confidence limits 6.7 - 11 mg/l.
- Other: None.

Reported statistics and error estimates:

None.

Validity criteria fulfilled:

yes

Conclusions:

The effect of the test material on the respiration of activated sewage sludge micro-organisms gave a 3-Hour EC50 of greater than 1000 mg/l. The No Observed Effect Concentration (NOEC) after 3 hours exposure was 1000 mg/l.

Executive summary:

Introduction.

A study was performed to assess the effect of the test material on the respiration of activated sewage sludge. The method followed that described in the OECD Guidelines for Testing of Chemicals (1984) No 209 "Activated Sludge, Respiration Inhibition Test", Method C.11 of Commission Regulation (EC) No. 440/2008 and US EPA Draft Ecological Effects Test Guidelines OPPTS 850.6800.

Methods. 

Activated sewage sludge was exposed to an aqueous dispersion of the test material at concentrations of 10, 32, 100, 320 and 1000 mg/l for a period of 3 hours at a temperature of approximately 21°C with the addition of a synthetic sewage as a respiratory substrate.

The rate of respiration was determined after 3 hours contact tiand compared to data for the control and a reference material, 3,5-dichlorophenol.

Results and Conclusion.

The effect of the test material on the respiration of activated sewage sludge gave a 3-Hour EC₅₀of greater than 1000 mg/l. The No Observed Effect Concentration (NOEC) after 3 hours exposure was1000 mg/l.

It was considered unnecessary and unrealistic to test at concentrations in excess of 1000 mg/l.

The reference material gave a 3-Hour EC₅₀value of 7.9 mg/l, 95% confidence limits 6.1 - 10 mg/l.

Description of key information

- (3-h) NOEC: ≥1000 mg/l; (3-h) EC50: > 1000 mg/l  [OECD 209; test mat. Distillates (Fischer–Tropsch), heavy, C18–C50 branched, cyclic and linear (nominal) based on: respiration rate]

- (3-h) NOEC: ≥1000 mg/l; (3-h) EC50: > 1000 mg/l  [OECD 209; test mat. Distillates (Fischer–Tropsch), C8–C26 branched and linear (nominal) based on: respiration rate]

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

1 000 mg/L

Additional information

The closely related substances GTL Gasoil (C8-C26) and GTL Base Oil Distillates (C18-C50) had no inhibitory effect upon activated sludge in tests conducted in accordance with OECD 209 and GLP. The 3 hour EC₅₀was >1000 mg/l; NOEC = 1000 mg/l (Clarke, 2009a and 2009b).

These data are used as read-across data to 'Paraffin waxes (Fischer-Tropsch), full-range, C15-50 - branched and linear'.