2H-Pyran, 2-[4'-[difluoro(3,4,5-trifluorophenoxy)methyl]-3',5'-difluoro[1,1'-biphenyl]-4-yl]-5-ethyltetrahydro-

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2008-01-28 to 2008-02-25

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1988-05-30

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Version / remarks:

1984-04-04

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

The test item is not soluble in water. Therefore, the test item was directly dosed into each test flask and tap water was added. For complete suspension of the test item, this mixture was stirred for about 24 hours at 21 °C in the dark / diffuse light before adding the inoculum. The test flasks were tightly closed to avoid water loss by evaporation.

Test organisms (species):

activated sludge, domestic

Details on inoculum:

Species / Origin:
Activated Sludge, micro organisms from a domestic waste water treatment plant was supplied by the municipal sewage treatment plant Darmstadt, Germany.

Conditioning:
The activated sludge was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was re-suspended in tap water and again centrifuged. The latter procedure was repeated twice. The washed sludge was resuspended in tap water and sewage feed (50 mL per L) was added. The sludge was kept at room temperature under continuous aeration until use over night. Immediately before use, the dry weight of the activated sludge was determined and diluted to 3.8 mg/L with tap water. The pH of the activated sludge was 7.8 and therefore no adjustment necessary.

Test type:

static

Water media type:

not specified

Limit test:

no

Total exposure duration:

3 h

Test temperature:

21°C during dissolution phase (24 hours); 19-21 °C during pre-incubation (3 hours); 18-20°C during evaluation period

pH:

At the start of the study the pH of the activated sludge was 7.8 and therefore no adjustment was necessary.
During the study pH values of 7.4 at the start and 8.6 at the end of study (after 3h) were determined.

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 8.5 mg O2/L and thus >6.5 mg O2/L.

Nominal and measured concentrations:

Nominal concentrations: 10, 32, 100, 320 and 1000 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass flasks of approximately 1 litre volume and Karlsruher flasks of 250 mL volume
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2
- No. of vessels per abiotic control (replicates): 1
- Sludge concentration (weight of dry solids per volume): 3.8 g/L

TEST MEDIUM / WATER PARAMETERS
- Source of dilution water: deionised water

Synthetic sewage:
16 g peptone
11 g meat extract
3 g urea
0.7g NaCl
0.4 g CaCl2 * 2H20
0.2 g MgSO4 * 7H2O
2.8 g K2HP04
filled up to 1 liter with deionised water

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: no detials available
- Light intensity: no detials available

EFFECT PARAMETERS MEASURED
respiration rate (Karlsruher flask)
The pH and the oxygen concentrations were determined at the start and at the end of the incubation period in all treatments. The water temperature was measured in one control medium at the start and the end of the incubation period.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: factor 3.2

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

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:

Based on measured inhibition rates, the 3 -hour EC10, EC20 and EC50 could not be quantified because up to the highest nominal test concentration of 1000 mg/L less than 10% inhibition was noted after three hours incubation. No dose related response was found. The test according to OECD 209 guideline does not provide a threshold value for the determination of a NOEC. If a deviation of less ≤ 15% between test item treated activated sludge and control is considered not to be an effect, the NOEC of the test item will be established above 1000 mg/L.

Validity Criteria of the Study
Inoculum Control: The respiration rates of the two controls did not differ by more than 15 %.
Reference Item: The 3-hour EC50 of the reference item 3,5-Dichlorophenol was determined to be 10.4 mg/L and is thus in the range of 5 to 30 mg/L for the used activated sludge batch.
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 8.5 mg O2/L and thus >6.5 mg O2/L.

Results with reference substance (positive control):

The 3 -hour EC50 of 3,5 -Dichlorophenol was calculated to be 10.4 mg/L with 95% confidence limits of 7.4 - 14.6 mg/L.

Reported statistics and error estimates:

The 3-hour EC50 including the 95 %-confidence limits of the reference item was calculated by Probit analysis. The 3-hour EC20 and EC50 of the test item could not be calculated due to the low toxicity of the test item to micro organisms under the tested conditions.

Validity criteria fulfilled:

yes

Conclusions:

The test according to OECD 209 guideline does not provide a threshold value for the determination of a EC50. Up to the highest nominal concentration of 1000 mg/L no respiration inhibition was detected (EC50 > 1000 mg/L, NOEC = 1000 mg/L).

Executive summary:

The influence of the test item on the activity of activated sludge was evaluated by measuring the respiration rate under defined conditions according to OECD 209 and EU Method C.11. The respiration rate (oxygen consumption) of an aerobic activated sludge fed with a standard amount of synthetic sewage was measured in the presence of various concentrations of the test item after an incubation period of 3 hours. The test concentrations were 10, 32,100, 320 and 1000 mg/L test item and 3.2,10 and 32 mg/L 3,5-Dichlorophenol. Additionally two inoculum controls were evaluated. The test item is not soluble in water. Therefore, the test item was directly weighted into the test vessels and stirred for about 24 hours before using in the experiment. The respiration rates of the activated sludge treated with the test item at concentrations between 10 mg/L and 1000 mg/L differed by less than 10% from control. Based on measured inhibition rates, the 3-hour EC10, EC20 and EC50 could not be quantified because up to the highest nominal test concentration of 1000 mg/L less than 10 % inhibition was noted after three hours incubation. No dose related response was found. The test according to OECD 209 guideline does not provide a threshold value for the determination of a NOEC. If a deviation of less < 15% between test item treated activated sludge and control is considered not be an effect, the NOEC of the test item will be established to be1000 mg/L. The 3-hour EC50 of 3,5-Dichlorophenol was calculated to be10.4 mg/L with 95 % confidence limits of 7.4 -14.6 mg/L. Up to the highest nominal concentration of 1000 mg/L no respiration inhibition was detected (EC50 > 1000 mg/L, NOEC = 1000 mg/L).

Description of key information

The test according to OECD 209 guideline does not provide a threshold value for the determination of a EC50. Up to the highest nominal concentration of 1000 mg/L no respiration inhibition was detected (EC50 > 1000 mg/L, NOEC = 1000 mg/L) (reference 6.1.7-1).

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

1 000 mg/L

Additional information

The influence of the test item on the activity of activated sludge was evaluated by measuring the respiration rate under defined conditions according to OECD 209 and EU Method C.11. The respiration rate (oxygen consumption) of an aerobic activated sludge fed with a standard amount of synthetic sewage was measured in the presence of various concentrations of the test item after an incubation period of 3 hours. The test concentrations were 10, 32,100, 320 and 1000 mg/L test item and 3.2,10 and 32 mg/L 3,5-Dichlorophenol. Additionally two inoculum controls were evaluated. The test item is not soluble in water. Therefore, the test item was directly weighted into the test vessels and stirred for about 24 hours before using in the experiment. The respiration rates of the activated sludge treated with the test item at concentrations between 10 mg/L and 1000 mg/L differed by less than 10% from control. Based on measured inhibition rates, the 3-hour EC10, EC20 and EC50 could not be quantified because up to the highest nominal test concentration of 1000 mg/L less than 10 % inhibition was noted after three hours incubation. No dose related response was found. The test according to OECD 209 guideline does not provide a threshold value for the determination of a NOEC. If a deviation of less < 15% between test item treated activated sludge and control is considered not be an effect, the NOEC of the test item will be established to be1000 mg/L. The 3-hour EC50 of 3,5-Dichlorophenol was calculated to be10.4 mg/L with 95 % confidence limits of 7.4 -14.6 mg/L. Up to the highest nominal concentration of 1000 mg/L no respiration inhibition was detected (EC50 > 1000 mg/L, NOEC = 1000 mg/L) (reference 5.1.7 -1).