Registration Dossier

Ecotoxicological information

Toxicity to microorganisms

Currently viewing:

Administrative data

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:
19/09/2011 - 19/03/2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP, Guideline study
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Version / remarks:
2010
Deviations:
yes
Remarks:
see Table A7.4.1.4-5
GLP compliance:
yes
Analytical monitoring:
no
Details on sampling:
At 30 min and 3 hours.
Vehicle:
no
Details on test solutions:
Not applicable
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
Nature: activated sludge
Species: not applicable
Strain: not applicable
Source: Municipal wastewater treatment plant, collected from the aeration tank
Sampling site: Pforzheim, Germany
Laboratory culture: No

Method of cultivation
Preparation of inoculum for exposure: The sludge was used one day after collection. It was settled for about 10 minutes and the upper layer with finer solids was decanted. Before starting the test, it was washed three times with chlorine free tap water by centrifugation (10 minutes at 3000 rpm). After centrifuging, the supernatant was decanted and discarded and the sludge was re-suspended in chlorine free tap water. This procedure was repeated twice.

The mixed liquor suspended solids (MLSS) were adjusted to a concentration of 3.0 g/L (± 10 %). The activated sludge was continuously aerated at the test temperature, the solids did not settle down.
Pretreatment: None
Initial cell concentration: Not reported
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Post exposure observation period:
none
Hardness:
Not reported
Test temperature:
17.6 – 20.0 °C (measured during O2-determination at 30 min and 3 h after start of respiration)
pH:
7.78 – 8.22
Dissolved oxygen:
Not reported
Salinity:
Not applicable
Nominal and measured concentrations:
Not reported
Details on test conditions:
Culturing apparatus: BOD flasks
Number of culture flasks/concentration:
- 3 replicates for the negative, the abiotic and the heterotrophic control (ATU)
- 3 replicates each at 1000mg/L for total oxygen uptake and heterotrophic oxygen uptake (ATU)
- 1 replicate each for the positive control (DCP), the 10 and 100 mg/L total oxygen uptake and heterotrophic uptake samples
Aeration device: air pump
Measuring equipment: O2-electrode: Dissolved Oxygen Hand-Held Meter OXI 340, WTW
Test performed in closed vessels due to significant volatility of TS: No
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol (DCP)
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Total inhibition, inhibition of the heterotrophic organisms and inhibition of nitrifying bacteria
Details on results:
NOEC = 1000 mg/L; EC50 > 1000 mg/L
Results with reference substance (positive control):
For the total oxygen uptake, the EC50 was determined to be 11.29 mg/L after 30 min and 6.17 mg/L after 3 h. These values were within the recommended range of 2 to 25mg/L. For the heterotrophic oxygen uptake, the EC50 was determined to be 23.03 after 30 min and 15.60 mg/L after 3h. These values were within the recommended range of 5 to 40 mg/L. For the oxygen uptake due to nitrification the EC50 was determined to be 2 mg/L after 30 min and 1.67 mg/L after 3h. Both values were within the recommended range of 0.1 to 10 mg/L
Reported statistics and error estimates:
The statistical evaluation was performed for specific respiration rate of the control and highest concentration using SAS® (2002–2008). The calculation was performed using SAS Software service pack 9.2. The NOEC was determined using the t-Test pooled. P-values below 0.05 showed statistically significant differences to the control. A test for normality of the data was performed by calculating the Shapiro-Wilk’s statistic. The EC50-values for the reference test item DCP were evaluated using probit analysis following normal and logistic distribution. In the case of the nitrifiers the EC50 value for DCP after 3 h was extrapolated with a linear regression curve. The EC50 calculations were based on the percentage of inhibition.

Table 1: Deviations from the study plan

Section of study plan

Deviation

Reason

Impact on the study

2.1 Test item

The date of the certificate of analysis changed from 05 September 2011 to 24 November 2010.

Mistake during study plan preparation.

None.

2.2 Reference item

The stock solution of the reference item was not prepared with 0.5 g with a final volume of 500 mL but with 0.25 g filled up to a final volume of 250 mL.

Mistake during study plan preparation

None, since the concentration was the same

2.5 Microbial Inoculum

The sludge was not used on the same day as collec-tion but on the next day.

Mistake in study performance

None, since the guideline allows storage up to two days

2.5 Microbial Inoculum

The sludge was not centrifuged for 20 min at 4500 rpm but for 10 min at 3000 rpm.

Mistake in study performance.

None, since the sludge was centrifuged to produce a clear supernatant and pellet of sewage solids, therefore fulfilling guideline criteria as the guide-line does not stipulate a speed of centrifugation.

2.8 Test conditions

The temperature of the test decreased to 17.6 °C.

Mistake in study performance

None, since the deviation did not have an adverse effect on the validity and outcome of the study. The guideline recommends a temperature range of 20 ± 2 °C. The lowest recorded temperature was 17.6°C. However, the test item showed no adverse effect and the EC50 value of the reference item was within the recommended range. Therefore, the temperature difference of 0.4°C is considered not to have had an adverse effect on the validity and the outcome of the study.

2.7.2 Procedure of the Test

The pH measured ranged from 7.92 – 8.22. This is above the recommended range of 7.5 ± 0.5.

Mistake in study performance

None, since the deviation did not have an adverse effect on the validity and outcome of the study. The test item showed no adverse effect and the EC50 of the reference item was within the recommended range. Therefore, the pH difference of 0.2 is considered not to have had an adverse effect on the validity and the outcome of the study.

 

Validity criteria fulfilled:
yes
Conclusions:
The test item dinotefuran technical had no toxic effects on the oxygen uptake at the highest concentration tested (1000 mg/L). Hence, the EC50 of dinotefuran technical is > 1000 mg/L.

Description of key information

The test item dinotefuran technical had no adverse effects on the oxygen uptake at the highest concentration tested (1000 mg/L). Hence, the EC50 of dinotefuran technical is > 1000 mg/L and the NOEC is 1000 mg/L.

Key value for chemical safety assessment

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

Additional information