ethyl 5,5-diphenyl-2-isoxazoline-3-carboxylate

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:

19 Nov 1999

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Deviations to current guideline: Only two control replicates and one replicate per exposure group were tested. Not all validity criteria are reported.

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Version / remarks:

adopted 1984

Qualifier:

according to guideline

Guideline:

other: Commission Directive 87/302/EEC: “Activated Sludge Respiration Inhibition Test”, Official Journal of the European Communities No. L 133/118-122, dated May 05, 1988.

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Preparation of inoculum for exposure: The sludge was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was resuspended in tap water and again centrifuged. The latter procedure was repeated twice. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge (g) to its dry weight (g) determined. Based on this ratio, calculated aliquots of washed sludge suspension, corresponding to 4 g dry material per litre, were made up with tap water. To this mixture, 50 mL synthetic sewage feed (see below) per litre was added daily, starting three days prior to use, and the sludge was kept at room temperature under continuous aeration until use. Immediately before use, the dry weight of the activated sludge was determined, and diluted to 4 g/L with tap water. The pH of the activated sludge was measured to be pH 7.3.

Test type:

static

Water media type:

freshwater

Total exposure duration:

3 h

Test temperature:

18 - 19 °C

pH:

Test start: 7.0 - 7.3
Test end: 8.1

Dissolved oxygen:

Test start: 7.5 - 8.3 mg/L
Test end: 6.8 - 7.6 mg/L

Nominal and measured concentrations:

Nominal: 10, 32, 100, 320, 1000 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: for incubation: glass flasks 1 L; for measurement of respiration: Karlsruher flasks 250 mL
- Material, fill volume: glass, 500 mL
- Aeration: compressed air at an air flow rate of 0.6 L/min
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2
- Biomass loading rate: 4 g dry sewage material per litre

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : The oxygen concentrations were determined before test start and at the end of the incubation period in all test concentrations and controls.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: approx. 3.2
- Test concentrations: 10, 32, 100, 320, 1000 mg/L

Reference substance (positive control):

yes

Remarks:

3,5-Dichlorophenol

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: EC50 was calculated to be 6 mg/L

In comparison to the inoculum controls, the respiration rate of the activated sludge was not inhibited (-0.5 to -19.0%) at the four lowest concentrations of nominal 10 to 320 mg/L. At the highest concentration of nominal 1000 mg/L, the respiration rate was moderately inhibited by 12.7%. Test substance concentrations exceeding 1000 mg/L nominal were not tested.

From these inhibition rates, the 3 h EC20 and the 3 h EC50 could not be quantified because up to the highest nominal test concentration of 1000 mg/L less than 20% inhibition was noted after three hours exposure time. Nevertheless, the 3 h EC20 and EC50 are far above 1000 mg/L nominal under the present test conditions.

Table 1: Validity criteria for OECD 209.

Criterion from the guideline	Outcome	Validity criterion fulfilled
The blank controls (without the test substance or reference substance) oxygen uptake rate should not be less than 20 mg oxygen per one gram of activated sludge (dry weight of suspended solids) in an hour.	Control 1 = 13.35 mg O2/ g * h Control 2 = 15 mg O2/ g * h	No
The coefficient of variation of oxygen uptake rate in control replicates should not be more than 30% at the end of definitive test.	< 30%	yes

Validity criteria fulfilled:

no

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Description of key information

EC50 (3 h) > 1000 mg/L (nominal, activated sludge, OECD 209)

Additional information

The respiration inhibition of the test item was tested in a respiration inhibition test using activated sludge fed with synthetic sewage (OECD 209). Only a moderate inhibition of 12.7% compared to the control was observed at the highest test item concentration of nominal 1000 mg/L. Thus, a 3 h EC50 for the test substance could not be quantified in this test and was reported as > 1000 mg/L.