Producto P0310

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

500.34 mg of the test item was weighed by means of an analytical balance and transferred to the designated test vessel with 284 mL tap water. The test item was dissolved in the tap water by intense stirring for fifteen minutes at room temperature. After stirring, the appearance of the test medium was evaluated, and the limit test concentration was a clear yellowish solution under the test conditions.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Preparation of inoculum for exposure: The sludge was washed twice with tap water by centrifugation and the supernatant liquid phase was decanted. A homogenized aliquot of the final sludge suspension was weighed, thereafter dried and the ratio of wet to dry weight was calculated. Based on this ratio, an aliquot of washed sludge was suspended in tap water to obtain a concentration equivalent to 3 g dry material per liter. During the holding period of three days prior to use, the sludge was fed daily with 50 mL synthetic wastewater per liter and was kept at room temperature under continuous aeration until use. Before use, the dry weight of the activated sludge was measured again in the inoculum used for the test.

The inoculum had a sludge concentration of 3.3 g/L dry weight (corresponding to about 1.3 g dry material per liter test medium).

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Test temperature:

20 ºC

pH:

7.6-8.5

Dissolved oxygen:

8.1-9.3 mg/L

Nominal and measured concentrations:

Nominal concentration: 1000 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 2000-mL glass beakers
- Aeration: the single test medium and the controls were continuously aerated by intense stirring on magnetic stirrers to avoid possible foaming and/or stripping of the test item.
- No. of vessels per concentration (replicates): one replicate
- No. of vessels per control (replicates): two replicates

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water
- Intervals of water quality measurement: The pH and dissolved oxygen concentrations were determined after the addition of synthetic wastewater and activated sludge in all test media and the controls at the start and at the end of the 3-hour incubation period. The water temperature was measured in one control at the start and at the end of the incubation period.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : For measurement of the respiration rate, a well-mixed sample of each test medium was poured into a BOD-flask after three hours incubation time, and was not further aerated. Then, the dissolved oxygen concentration was measured with an oxygen electrode (WTW TriOxmatic® 300 and an oxygen meter WTW Oxi 539, Wissenschaftlich-Technische Werkstaetten WTW, Weilheim/Germany), and was continuously recorded. During measurement, the samples were continuously stirred on a magnetic stirrer. The oxygen consumption rate (in mg O2 L-1 minute-1) was determined from the linear part of the respiration curve in the range 6.5–2.5 mg O2/L. In case of very rapid oxygen consumption, the range used was below the limits indicated above, but always within the linear part of the respiration curve. In case of low oxygen consumption, the rate was determined over a period of at least ten minutes.

Reference substance (positive control):

yes

Remarks:

3,5-Dichlorophenol

Key result

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Details on results:

The test item had no significant inhibitory effect (<15%) on the respiration rate of activated sludge after the incubation period of three hours at the limit test concentration of 1000 mg/L. Based on the physical appearance, the test solution was a clear, yellowish solution.

Results with reference substance (positive control):

The 3-hour EC50 of the reference item 3,5-dichlorophenol (positive control) was calculated to be 20 mg/L (the 95% confidence limits could not be calculated). The 3-hour EC50 is within the guideline-recommended range of 5–30 mg/L, confirming suitability of the activated sludge used.

Validity criteria fulfilled:

yes

Remarks:

(The oxygen consumption rates of the two controls differed by 7% (guideline-recommended maximum variation: 15%; results obtained with the positive control are within the acceptable range).

Conclusions:

The 3-hour NOEC (EC15) of the test item to activated sludge microorganisms was at least 1000 mg/L.

Executive summary:

The inhibitory effect of the test item on the respiration rate of aerobic wastewater microorganisms of activated sludge was investigated in a 3-hour respiration inhibition test according to the EU Commission Directive 88/302/EEC, Part C.11, and the OECD Guideline for Testing of Chemicals, No. 209. In accordance with the EU Commission Directive 92/69/EEC (C.1, C.2, and C.3), a limit test with a nominal test concentration of 1000 mg/L was performed. In addition, two controls and three different concentrations of the reference item 3,5-dichlorophenol (5, 16, and 50 mg/L) were tested in parallel. The results of these treatments confirmed suitability of the activated sludge and the method used. The test item had no inhibitory effect on the respiration rate of activated sludge after the incubation period of three hours at the test item concentration of 1000 mg/L. Thus, the 3-hour NOEC (EC15) of the test item to activated sludge microorganisms was at least 1000 mg/L. This value might even be higher, but concentrations above 1000 mg/L were not tested. The 3-hour EC20, EC50, and EC80 could not be calculated, but were clearly higher than 1000 mg/L.

Description of key information

Key study: OECD guideline 209 and EU method C.11. GLP study.

The 3-hour NOEC (EC15) of the test item to activated sludge microorganisms was at least 1000 mg/L.

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

1 000 mg/L

Additional information

Key study: OECD guideline 209 and EU method C.11. GLP study.

The 3-hour NOEC (EC15) of the test item to activated sludge microorganisms was at least 1000 mg/L.