Disodium ar,ar'-[(9,10-dihydro-9,10-dioxo-1,4-anthrylene)diimino]bis[(4-chlorophenoxy)benzenesulphonate]

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:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Version / remarks:

adopted ApriI 4, 1984

GLP compliance:

no

Details on test solutions:

The test article was dissolved in bidistilled water at a concentration of 500 mg/l.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Source: the tests were run with aerobic activated sludge from a domestic waste-water treatment plant (ARA, Sissach/Switzerland).
- Pretreatment: the sludge was coarsely sieved, centrifuged, the supernatant liquid phase decanted, the solid material resuspended in tap water and again centrifuged. This procedure was repeated a further two times.
- Initial biomass concentration: a volume of washed sludge suspension corresponding to 4.0 g dry material were made up with Soerensen buffer (pH = 7) to 1 litre.
- Preparation of inoculum for exposure: 50 ml OECD recommended synthetic sewage feed were added to sludge suspension.
- Incubation: the sludge was incubated overnight at 20 - 25 °C under aeration.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

pH:

7.8 at the start

Nominal and measured concentrations:

10 and 100 mg/l nominal

Details on test conditions:

TEST SYSTEM
- Test vessel: glass-flasks (> 500 ml).
- Aeration: 0.2 l/minute instead of 0.5 l/minute to prevent possible foam development.
- No. of vessels per concentration: one replicate.
- No. of vessels per control: 2 replicates.
- Sludge amount: 200 ml

SAMPLING AND ANALYSIS
After 180 minutes of aeration at 22.0 °C, the first sample was taken for oxygen measurement over a period of up to 10 minutes. Thereafter, in 15 minutes intervals, the rest of the samples were submitted to measurement of oxygen concentration.
Sampling was started with the control 1. Thereafter, the measurements were continued with the positive controls and the test article samples. Finally, the oxygen consumption of control 2 was determined.

MEASUREMENTS
Oxygen consumption was measured over a period of 10 minutes. During measurement, the sample was not aerated but continuously stirred on a magnetic stirrer.

VALIDITY OF THE TEST
The tests are validated by testing a known standard, i.e. 3,5-dichlorophenol. The IC50-value should be in the range of 5 - 30 mg/l. Furthermore, the variation between the two control samples (start, end) should not be greater than 15 %.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol, 3.2, 10 and 50 mg/l

Duration:

3 h

Dose descriptor:

IC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Details on results:

The test article showed only a slight inhibition (17.0 %) of the respiration rate up to a concentration of 100 mg/l.
The aeration of the test medium with test article caused the solutions to foam strongly.

Results with reference substance (positive control):

At a concentration of the standard of 10 mg/l, the respiration rate was highly inhibited by 75.5 %. The IC50 was found to be about 9.0 mg/l. Almost complete inhibition (91.5 %) was observed at 50 mg/l.

Influence of test item and reference substance on oxygen consumption of activated sludge.

Test substance (mg/l)	Oxygen consumption (mgO2/l min)	Inhibition (%)	Starting pH
Control	0.88	-	7.8
Control	1.00	-	7.8
Mean	0.94± 0.06 (13.6 %)
Test item 100	0.78	17.0	7.8
Test item 10	0.91	3.2	7.8
3 ,5-dichlorophenol 50	0.08	91.5	7.8
3 ,5-dichlorophenol 10	0.23	75.5	7.8
3 ,5-dichlorophenol 3.2	0.88	6.4	7.8

Validity criteria fulfilled:

yes

Remarks:

the IC50-value of reference substance was in the range of 5 - 30 mg/l and the variation between the two control samples (start, end) was not greater than 15 %

Conclusions:

IC50 (3h) > 100 mg/l (nominal)

Executive summary:

The inhibitory effect of test item on aerobic waste-water bacteria was investigated in a respiration test, according to the OECD guideline 209. The method determines the respiration rate (oxygen consumption) of an aerobic activated sludge fed with an excess but standard amount of biodegradable food and as compared to the respiration rate obtained in an activated sludge run under identica conditions but containing the test chemical.

The test article showed only a slight inhibition (17.0 %) of the respiration rate up to a concentration of 100 mg/l. The IC50 was determined to be higher than 100 mg/l.

Conclusion

IC50 (3h) > 100 mg/l (nominal)

Description of key information

IC50 (3h) > 100 mg/l (nominal)

Key value for chemical safety assessment

EC50 for microorganisms:

100 mg/L

Additional information

The inhibitory effect of test item on aerobic waste-water bacteria was investigated in a respiration test, according to the OECD guideline 209. The test article showed only a slight inhibition (17.0 %) of the respiration rate up to a concentration of 100 mg/l. The IC50 was determined to be higher than 100 mg/l.