Oxazolidine, 3-butyl-2-(1-ethylpentyl)-

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

Study period:

from 2005-07-12 to 2005-07-21

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

Version / remarks:

adopted 04.11.1984

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted 1988

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.6800 (Modified Activated Sludge, Respiration Inhibition Test for Sparingly Soluble Chemicals)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Vehicle:

no

Details on test solutions:

For the purpose of the definitive test, the test material was dispersed directly in water.
Amounts of test material (90, 160, 280, 500, 900 and 1600 mg) were each separately dispersed in approximately 250 mL of water and subjected to ultrasonication for approximately 30 minutes. Synthetic sewage (16 mL), activated sewage sludge (200 mL) and water were added to a final volume of 500 mL to give the required concentrations of 180, 320, 560, 1000, 1800 and 3200 mg/L.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

The activated sewage sludge sample was maintained on continuous aeration in the laboratory at a temperature of 21 °C and was used on the day of collection. The pH of the sample was 7.9 measured using a WTW pH/Oxi 3401 pH and dissolved oxygen meter. Determination of the suspended solids level of the activated sewage sludge was carried out by filtering a sample (100 mL) of the activated sewage sludge by suction through a pre-weighed GF/A filter paper using a Büchner funnel which was then rinsed 3 times with 10 mL of deionised reverse osmosis water and filtration continued for 3 minutes. The filter paper was then dried in an oven at approximately 105 °C for at least 1 hour and allowed to cool before weighing. This process was repeated until a constant weight was attained. The suspended solids were equal to 3.8 g/L prior to use.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Hardness:

approximately 100 mg/L as CaCO3.

Test temperature:

21 °C

pH:

Control: 8.1 - 8.2

Test item:
180 mg/L: 8.3
320 mg/L: 8.4
560 mg/L: 8.7
1000 mg/L 8.9
1800 mg/L: 9.1
3200 mg/L: 9.2

3,5-dichlorophenol
3.2 mg/L: 8.3
10 mg/L: 8.4
32 mg/L: 8.5
3200 mg/L: 9.2

Dissolved oxygen:

approximately 6.3 - 7.2 mg O2/L

Nominal and measured concentrations:

Nominal concentrations 180, 320, 560, 1000, 1800 and 3200 mg/L.
No measured concentrations.

Details on test conditions:

TEST SYSTEM
- Test vessel: 500 mL conical flask
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: dechlorinated tap water
- Total organic carbon: average 1.675 mg/L
- Total pesticides: below LOD
- Total chlorine: average 0.153 mg/L
- Sodium: 24.625 mg/L

OTHER TEST CONDITIONS
- Photoperiod: normal laboratory lighting

EFFECT PARAMETERS MEASURED:
respiration rate

TEST CONCENTRATIONS
Initial Range-finding test:
100 and 1000 mg/L.
Range-finding test:
1.0, 10 and 90 mg/L
Second Range-finding test
100 and 1000 mg/L
Definitive test:
180, 320, 560, 1000, 1800, and 3200 mg/L

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Duration:

30 min

Dose descriptor:

EC50

Effect conc.:

> 3 200 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

1 400 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Key result

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

180 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

The validation criteria for the control respiration rates and reference material EC50 values have been satisfied.
In some instances, the initial and final dissolved oxygen concentrations were below those recommended in the test guidelines (6.5 mg O2/L and 2.5 mg O2/L, respectively). This was considered to have had no adverse effect on the results of the study given that in all cases the oxygen consumption rate was determined over the linear portion of the oxygen consumption trace.
Observations made throughout the definitive test, showed that at all of the test concentrations, an oily slick of test material was visible on the surface of the test media. This was considered to be due to the insoluble nature of the test material in the test medium. However, the test material was dispersed with the aid of ultrasonication in the test diluent for approximately 30 minutes prior to the addition of the synthetic sewage, activated sewage sludge and water (to a final volume of 500 mL) and each vessel was aerated with compressed air at a rate of approximately 0.5 - 1 litre per minute to ensure that there was maximum contact between the test material and activated sewage sludge in the test medium.

Relatively low initial oxygen readings were observed in the test vessels which could possibly have been due to insoluble material coating the membrane of the oxygen probe. This was considered not to have had a significant effect on the results of the test as the oxygen consumption rate was determined over the linear portion of the oxygen consumption trace.
The test material vessels showed a concentration dependent increase in pH values with the higher test concentrations having higher pH values. This may have been a contributing factor to the toxic nature of the test material but it was considered not to affect the integrity of the study given that the purpose of the study was to determine the effect that the test material would have on a waste water treatment facility where pH adjustment of the incoming effluent would not occur.

Results with reference substance (positive control):

3,5-dichlorophenol
EC50 (3 hours) 7.1 mg/L

Inhibition values (%) after 3 h:
3.2 mg/L: 33%
10 mg/L: 57%
32 mg/L: 83%

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the performed Activated Sludge Respiration Inhibition Test, the EC50 value for the test item following 3 h incubation was determined as 1400 mg/L. The No Observed Effect Concentration (NOEC) after 3 hours was determined as 180 mg/L.

Executive summary:

The purpose of the 3-hour test was to evaluate the influence of the test item on the activity of activated sludge by measuring the respiration rate under defined conditions. The test was performed in accordance to the OECD guideline No 209 and to USEPA method OPPTS 850.6800. The respiration rates of activated sludge exposed to the test item were measured after 30 min and after 3 h by using a Yellow Springs dissolved oxygen meter fitted with a BOD probe.

Following initial range finding studies, the nominal concentrations used for the definitive test were 180, 320, 560, 1000, 1800, and 3200 mg/L. The negative control group was maintained under identical conditions but not exposed to the test material. A reference material, 3,5-dichlorophenol, was included in the range-finding test at concentrations of 3.2 and 32 mg/L in order to confirm the suitability of the inoculum. The following results (nominal values) were obtained for the test item: EC50 (30 minutes): >3200 mg/L, EC50 (3 hours): 1400 mg/L. The No Observed Effect Concentration (NOEC) after 3 hours was 180 mg/L.

Description of key information

Under the conditions of the performed Activated Sludge Respiration Inhibition Test, the EC50 value for the test item following 3 h incubation was determined as 1400 mg/L. The No Observed Effect Concentration (NOEC) after 3 hours was determined as 180 mg/L.

Key value for chemical safety assessment

EC50 for microorganisms:

1 400 mg/L

EC10 or NOEC for microorganisms:

180 mg/L

Additional information

The purpose of the 3-hour test was to evaluate the influence of the test item on the activity of activated sludge by measuring the respiration rate under defined conditions. The test was performed in accordance to the OECD guideline No 209 and to USEPA method OPPTS 850.6800. The respiration rates of activated sludge exposed to the test item were measured after 30 min and after 3 h by using a Yellow Springs dissolved oxygen meter fitted with a BOD probe.

Following initial range finding studies, the nominal concentrations used for the definitive test were 180, 320, 560, 1000, 1800, and 3200 mg/L. The negative control group was maintained under identical conditions but not exposed to the test material. A reference material, 3,5-dichlorophenol, was included in the range-finding test at concentrations of 3.2 and 32 mg/L in order to confirm the suitability of the inoculum. The following results (nominal values) were obtained for the test item: EC50 (30 minutes): >3200 mg/L, EC50 (3 hours): 1400 mg/L. The No Observed Effect Concentration (NOEC) after 3 hours was 180 mg/L.