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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:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Qualifier:
according to
Guideline:
other: EEC Directive 67/548, 1987 (87/302), Part C: Methods for the determination of ecotoxicity, Publication No. L133, "Biodegradation: Activated sludge respiration inhibition test".
GLP compliance:
yes
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution of 0.5 g/L was prepared in Milli-Q-water. The pH of the stock solution was 8.5. Five concentrations were tested: 32, 56, 100, 180 and 284 mg/L, corresponding to 32, 56, 100, 180 and 284 of the prepared stock solution in 500 mL final volume.
Test organisms (species):
activated sludge
Details on inoculum:
- Source: Municipal sewage treatment plant: Waterschap de Maaskant', 's-Hertogenbosch, the Netherlands
- Preparation of inoculum for exposure: The sludge was coarsely sieved and wshed with tap-water. A small amount of the sludge was weighed and dreied at 105 °C to determine the amount of suspended solids (2.5 g/L of sludge). The batch sludge was used on subsequent days (maximum four days), therefore 50 mL of synthetic sewage was added to each litre of activated sludge at the end of each working day. The sludge was kept aerated at test temperature until use. Before use the PH was checked (measured value: 7.7)
- composition of the synthetic sewage feed: 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 l Milli-Q-water and filtered; the pH was 7.0
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
30 min
Test temperature:
20 °C
pH:
7.0-7.4
Nominal and measured concentrations:
nominal concentrations: 32, 56, 100, 180 and 284 mg/L
Details on test conditions:
TEST SYSTEM
- Type: closed
- Material, size, headspace, fill volume: Glass, 500 mL beakers and 300 mL oxygen bottles
- Aeration: whole test period except during measurements
- No. of organisms per vessel: 200 mL activated sludge
- No. of vessels per concentration (replicates): 1 (test substance) and 1 (reference substance)
- No. of vessels per control (replicates): 2

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Tap-water purified by reverse osmosis and subsequently passed over activated carbon and ion-exchange cartridges (Millipore Corp., Bedfored, Mass., USA)

OTHER TEST CONDITIONS
- stirring with magnetic stirrer during the wohle test period including measurements

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
- Oxygen consumption was measured and recorded for approximately 10 min. (Oxygen electrode Tri Ox EO 200, WTW, FRG supplied with a recorder Kipp BD40


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1. -1.8
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Duration:
30 min
Dose descriptor:
EC50
Effect conc.:
> 284 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Details on results:
The mean respiration rate of the control 1 and 2 was 37 mg O2/l/hr. The differences between the controls was 3%. The mean respiration rate of the control 2 and 3 was 36 mg O2/L/hr. The differences between the controls was 0%.
Since all criteria for acceptability of the test were met, this study was considered to be valid.
Results with reference substance (positive control):
The EC50 of 3,5-dichlorophenol was 7 mg/L (regression line: Y = 54.9282 x +4.8887)

Table2: The oxygen concentration at the start of the measurements and the influence of 3,5-dichlorophenol and A-1160 (methanol stripped) on the oxygen consumption of microbes in activated sludge.

Flask

Concentration reference/test substance (mg/L)

Oxygen conc. At the start of measurement (- mg O2/L)

Oxygen consumption (mg O2/L/hr)

Inhibition %

pH

C1

-

8.4

37

-

7.3

C2

-

8.3

36

-

7.1

Mean C1 + C2

37 (¿3%)

 

R1

3.2

8.6

26

29

7.2

R2

10

9.0

12

67

7.2

R3

32

8.9

6

84

7.4

C2

-

8.3

36

-

7.1

C3

-

8.3

36

-

7.0

Mean C2 + C3

36 (¿0%)

 

T1

32

8.5

37

-31

7.1

T2

56

8.1

39

-81

7.1

T3

100

8.2

40

-117

7.1

T4

180

7.3

37

-37

7.1

T5

284

8.2

44

-221

7.2

C: Control.

R: Reference substance, 3,5-dichlorophenol.

T: Test substance, A-1160 (methanol stripped).

1: A negative value indicate stimulation in respiration rate of the sludge.

Conclusions:
Under the condtion of this present test, A-1160 (methanol stripped) was not toxic to waste-water (activated sludge) bacteria at concentrations at or below 284 mg/L. Hence, the EC50 for A-1160 (methanol stripped) exceeded 284 mg/L.
Endpoint:
activated sludge respiration inhibition testing
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
Please refer to the attached justification below and the overall justification for grouping of substances attached in IUCLID Section 13.
Reason / purpose:
read-across source
Duration:
30 min
Dose descriptor:
EC50
Effect conc.:
> 284 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate

Description of key information

EC50 (30 min) > 284 mg/L (nominal, OECD 209) based on read across from CAS 116912-64-2

Key value for chemical safety assessment

Additional information

No data are available testing the toxicity of [3-(trimethoxysilyl)propyl]urea (CAS 23843-64-3) to microorganisms, therefore good quality data from the analogue substance, ureidopropyltrialkoxysilane, mixed methoxy and ethoxy esters (CAS 116912-64-2) have been read across. Both substances share the same silanol hydrolysis product (i.e. [3-(trihydroxysilyl)propyl]urea). The other hydrolysis products are methanol and ethanol. Details on read across justifications can be found in the attached justification in the respective target entry and in the overall justification for grouping of substances attached in IUCLID Section 13.

 

The influence of the test item ureidopropyltrialkoxysilane, mixed methoxy and ethoxy esters (CAS 116912-64-2) on the respiration rate of activated sludge was investigated according to OECD 209 and GLP (1999). The nominal test concentrations were 32, 56, 100, 180, 284 mg/L. No inhibition on respiration rate of the sludge was recorded in any of the concentrations tested. Therefore, the EC50 for ureidopropyltrialkoxysilane, mixed methoxy and ethoxy esters (CAS 116912-64-2) is > 284 mg/L. The respiration rates of the control were within 15% of each other. The EC50 of the reference substance, 3,5 -dichlorophenol, was 7 mg/L. Therefore, the test was considered to be valid.

The non-silanol hydrolysis products methanol and ethanol are well characterised in the public domain literature and are not hazardous at the concentrations relevant to the studies (OECD 2004a, OECD 2004b).

 

References:

OECD (2004a): SIDS Initial Assessment Report for SIAM 19, Berlin, Germany, 18-20 October 2004, Methanol, CAS 67-56-1.

OECD (2004b): SIDS Initial Assessment Report for SIAM 19, Berlin, Germany, 19-22 October 2004, Ethanol, CAS 64-17-5.