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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:
2009-11-12 to 2009-12-16
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
The study was conducted according to an appropriate OECD test guideline, and in compliance with GLP, and was considered to be reliability 1 (reliable without restrictions).
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:
yes
Analytical monitoring:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Known volume of test substance, Milli-RO water, synthetic sewage feed (16 ml) and activated sludge (200 ml) were mixed and made up to 500 ml with Milli-RO water in a 1 litre bottle.
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
- Preparation of inoculum for exposure: Coarsely sieved then washed by centrifuging at 1000 g for 10 minutes and supernatant decanted and replced with ISO-medium. Washing step performed three times.

- Amount suspended solids: 4.0 g/l
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Post exposure observation period:
Oxygen consumption measured and recorded for approximately 10 minutes.
Test temperature:
Experiment 1: between 19.4 and 20.0 °C; Experiment 2: between 18.5 and 19.4°C.
pH:
Experiment 1: between 6.8 and 6.9; Experiment 2: between 2.0 and 7.2.
Dissolved oxygen:
Oxygen concentration at start (=~ mg O2/l). Experiment 1: 6.2 to 6.7; Experiment 2: 7.7 to 9.6.
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: All glass, approximately 300 ml oxygen bottles and 1 litre test bottles.
- Aeration: Strong aeration with clean, oil-free air.

- No. of vessels per concentration (replicates): Experiment 1: Loading rate tested in duplicate. Experiment 2: Five loading rates.

- No. of vessels per control (replicates): Experiment 1 and 2. Control vessels in duplicate at beginning and end of each series (test substance and reference substance).

- No. of vessels per reference (replicates): Experiment 1 and 2. Three concentrations.


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

OTHER TEST CONDITIONS
- Adjustment of pH: None

Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
160 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.:
32 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:
EC50
Effect conc.:
ca. 83.5 mg/L
Conc. based on:
other: registered substance
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Duration:
3 h
Dose descriptor:
NOEC
Effect conc.:
ca. 16.7 mg/L
Nominal / measured:
nominal
Conc. based on:
other: registered substance
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Details on results:
25% inhibition of respiration rate of the sludge was recorded at 100 mg/l, therefore a second experiment was performed with a range of loading rates (10 to 1000 mg/l). The inhibition at a loading rate of 100 mg/l was slightly lower in the second experiment than in the first experiment. No significant inhibition was recorded at and below a loading rate of 32 mg/l. Almost complete inhibition was recorded at and above a loading rate of 320 mg/l. It is likely that the inhibition was influenced by the low pH levels at the higher loading rates, which dropped to pH 3.5 and pH 2.0 at 320 and 1000 mg/l respectively (pH at 100 mg/l was 6.8 - 6.9 in both experiments).
Results with reference substance (positive control):
- Results with reference substance valid? Yes. The 3-hr EC50 was in the accepted range of 5-30 mg/l (7.7 mg/l in Experiment 1; 6.1 mg/l in Experiment 2.)

Table 1: Experiment 1. Controls (C) and Dichloro(methyl)silane (T): pH, oxygen concentration, oxygen consumption and percentage inhibition of the respiration rate.

Flask

Loading rate (mg/l)

Oxygen conc. At the start (=~ mg/O2/l)

Oxygen consumption

(mg O2/l/h)

% Inhibition

respiration rate

pH

Cstart T

0

6.9

60

-

7.5

Cend T

0

6.4

68

-

7.4

 

∆13%

 

 

T1

100

6.2

48

25

6.9

T2

100

6.7

48

25

6.8

 

Table 2: Experiment 2. Controls (C) and Dichloro(methyl)silane (T): pH, oxygen concentration, oxygen consumption and percentage inhibition of the respiration rate.

Flask

Loading rate (mg/l)

Oxygen conc. At the start (=~ mg/O2/l)

Oxygen consumption

(mg O2/l/h)

% Inhibition

respiration rate

pH

Cstart T

0

8.1

40

-

7.2

Cend T

0

7.9

43

-

7.1

 

∆8%

 

 

T1

10

7.7

41

1

7.2

T2

32

7.7

39

6

7.0

T3

100

7.8

37

11

6.8

T4

320

9.3

1

98

3.5

T5

1000

9.6

0

100

2.0

 

 

Validity criteria fulfilled:
yes
Conclusions:
Read-across from dichloromethylsilane to the registered substance is justified because this substance rapidly hydrolyses to methylsilanediol and ultimately methylsilanetriol, which are the same as the hydrolysis products from the registered substance.
An EC50 of 160 mg/l and a NOEC of 32 mg/l for toxicity to microorganisms were determined in a reliable study conducted according to an appropriate test protocol, and in compliance with GLP. It is likely that the inhibition was influenced by the low pH levels at the higher loading rates, but the pH at the NOEC was 7, and therefore the NOEC is considered the most useful and representative result.


Description of key information

Toxicity to microorganisms: ASRI NOEC 32 mg/l (OECD 209), based on read-across from a structurally-related substance.

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:
32 mg/L

Additional information

No data are available for the submission substance, HD4

However, the substance is predicted to hydrolyse rapidly to methylsilanediol, which is then expected to hydrolyse further to methylsilanetriol.

Therefore, good quality data for the structurally-related substance dichloro(methyl)silane (CAS 75-54-7), have been read across.

Dichloro(methyl)silane and methylsilanediol are within an analogue group of substances that exhibit some evidence of toxicity to microorganisms. Dichloro(methyl)silane hydrolyses very rapidly to methylsilanediol.

The ultimate hydrolysis product of the registration substance (methylsilanetriol) is not within this analogue group, as it does not have a Si-H bond. Methylsilanetriol is within an analogue group of substances that exhibit no evidence of toxicity to microorganisms. The intermediate hydrolysis product of the registration substance (methylsilanediol) is more relevant to the WWTP exposure than the triol, given the typical residence time in a WWTP and represents a worst-case for toxicity to microoganisms.

It is therefore considered valid to read-across the results for dichloro(methyl)silane, as a worse case, to fill the data gap for the registered substance.

Additional information is given in a supporting report (PFA, 2013k) attached in Section 13 of the IUCLID dossier.

An ASRI EC50 of 160 mg/l and a NOEC of 32 mg/l for toxicity to microorganisms were determined for dichloro(methyl)silane. It is likely that the inhibition was influenced by the low pH levels at the higher loading rates, but the pH at the NOEC was 7, and therefore the NOEC is considered the most useful and representative result.

The study is considered to be reliability 1 (reliable without restrictions).