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Toxicity to microorganisms

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Reference
Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
09 February 2012 to 02 March 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline 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:
ISO 8192 (Water quality - Test for inhibition of oxygen consumption by activated sludge for carbonaceous and ammonium oxidation)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
The batch of Triethoxy(3-thiocyanatopropyl)silane tested was a clear light brown liquid with a purity of 98.3%.

The test substance was pipetted below the surface of the Milli-RO-water and stirred briefly to mix (5 minutes). Subsequently, 16 ml synthetic sewage feed, 250 ml sludge and Milli-RO water up to 500 ml were added resulting in the required loading rates. Optimal contact between the test substance and test organisms was ensured applying continuous aeration and stirring.
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
The sludge was coarsely sieved, washed and diluted with ISO-medium. A small amount of the sludge was weighed and dried overnight at ca. 105°C to determine the amount of suspended solids (3.0 g/l of sludge, as used for the test). The pH was 7.8 on the day of testing. The batch of sludge was used one day after collection; therefore 50 ml of synthetic sewage feed was added per litre of activated sludge at the end of the collection day. The sludge was kept aerated at test temperature until use.
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
3 h
Post exposure observation period:
After the 3-hour contact time, the oxygen consumption was recorded for a period of approximately 10 minutes. During measurement, the sample was not aerated but continuously stirred on a magnetic stirrer. The pH and temperature were determined in the remaining part of the reaction mixture.

Test temperature:
Between 18.2 and 20.1°C
pH:
Before addition of sludge: 7.6 - 7.8
After the 3 hour exposure period: 7.9 - 8.3
Dissolved oxygen:
The oxygen concentration at the start of measurements was at least 7.2 mg O2/l

Oxygen consumption:
Controls: 42 mg O2/l h
Triethoxy(3-thiocyanatopropyl)silane 1.0 mg/l: 33 mg O2/l h
Triethoxy(3-thiocyanatopropyl)silane 3.2 mg/l: 35 mg O2/l h
Triethoxy(3-thiocyanatopropyl)silane 10 mg/l: 33 mg O2/l h
Triethoxy(3-thiocyanatopropyl)silane 32 mg/l: 28 mg O2/l h
Triethoxy(3-thiocyanatopropyl)silane 100 mg/l: 23 mg O2/l h
Triethoxy(3-thiocyanatopropyl)silane 320 mg/l: 19 mg O2/l h
Triethoxy(3-thiocyanatopropyl)silane 1000 mg/l: 10 mg O2/l h
Nominal and measured concentrations:
Nominal loading rates: 1.0, 3.2, 10, 32, 100, 320 and 1000 mg/l.
Details on test conditions:
TEST SYSTEM
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: All glass
- Aeration:During exposure with clean, oil-free air
- No. of vessels per concentration (replicates): 5
- No. of vessels per control (replicates): 6 series
- Biomass loading rate: 1.5 g/l suspended solids in final test mixture

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Tap-water purified by reverse osmosis (Milli-RO
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : The respiration rate from each vessel, in mg O2/l/hr, was calculated from the linear part of the respiration curve, which was generally between 2 and 7 mg O2/l.

TEST CONCENTRATIONS
- Test concentrations: full test at 1.0, 3.2, 10, 32, 100, 320 and 1000 mg/l (loading rates)
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Key result
Duration:
3 h
Dose descriptor:
NOEC
Effect conc.:
< 1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
(loading rate)
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Key result
Duration:
3 h
Dose descriptor:
EC10
Effect conc.:
4.3 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
(loading rate)
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Remarks on result:
other: (2.0-9.3)
Key result
Duration:
3 h
Dose descriptor:
other: EC20
Effect conc.:
10 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
(loading rate)
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Remarks on result:
other: (4.8 - 21)
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
130 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
(loading rate)
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Remarks on result:
other: (68 - 260)
Key result
Duration:
3 h
Dose descriptor:
other: EC80
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
(loading rate)
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Details on results:
Statistically significant inhibition of the respiration rate of the sludge was recorded at all loading rates tested (Bonferroni t-Test: α=0.05 Toxstat).
The results of this test allowed for reliable determination of EC values. The NOEC was below the lowest loading rate tested. Testing at lower concentration is not possible for practical reasons.
Results with reference substance (positive control):
The EC50 of 3,5-dichlorophenol was in the accepted range of 2 to 25 mg/l for total respiration (5.6 mg/l in the final test).
Reported statistics and error estimates:
ECx
The percentage inhibition was plotted against the logarithm of the loading rates. The EC50, and if possible the EC10, EC20 and EC80 for the test substance, were determined using linear regression analysis. If available, the 95% confidence limits of the EC values were given.

NOEC estimation
In this present test Triethoxy(3-thiocyanatopropyl)silane was toxic to waste water (activated sludge) bacteria at all loading rates tested. Thus, the NOEC was below the lowest loading rate tested (NOEC < 1.0 mg/l). Testing at lower concentration is not possible for practical reasons.

Validity criteria fulfilled:
yes
Conclusions:
Under the conditions of this present test, triethoxy(3-thiocyanatopropyl)silane was toxic to waste water (activated sludge) bacteria at all loading rates tested (NOEC < 1.0 mg/l).

The EC10 was at a loading rate of 4.3 mg/l (95% confidence interval: 2.0 – 9.3 mg/l).
The EC20 was at a loading rate of 10 mg/l (95% confidence interval: 4.8 - 21 mg/l).
The EC50 was at a loading rate of 130 mg/l (95% confidence interval: 68 – 260 mg/l).
The EC80 was above the highest loading rate tested (> 1000 mg/l).

Description of key information

Toxicity to microorganisms:  3-hour ASRI EC50 130 mg/L (68 - 260 mg/L); EC10 4.3 mg/L (2.0 – 9.3 mg/L); NOEC <1 mg/L (OECD 209)

Key value for chemical safety assessment

EC50 for microorganisms:
130 mg/L
EC10 or NOEC for microorganisms:
4.3 mg/L

Additional information

The toxicity of the registered substance to microorganisms has been investigated in accordance to OECD Test Guideline 209 and in compliance with GLP. Under the conditions of this present test triethoxy(3-thiocyanatopropyl)silane was toxic to wastewater (activated sludge) bacteria at all loading rates tested (NOEC < 1.0 mg/l).

The EC10 was at a loading rate of 4.3 mg/L (95% confidence interval: 2.0 – 9.3 mg/L).

The EC20 was at a loading rate of 10 mg/L (95% confidence interval: 4.8 - 21 mg/L).

The EC50 was at a loading rate of 130 mg/L (95% confidence interval: 68 – 260 mg/L).

The EC80 was above the highest loading rate tested (> 1000 mg/L).