cyclohexadecanone

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:

16 Feb 2001

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:

Apr 1984

Deviations:

yes

Remarks:

1) Copper (II) sulphate pentahydrate was used instead of 3,5-dichlorophenol. 2) A dry sludge concentration of 3 g/L ± 20 % was used instead of 4 g/L ± 10 %. These deviations have no impact on the quality of the study.

GLP compliance:

yes (incl. QA statement)

Remarks:

Niedersächsisches Umweltministerium, Hannover, Germany

Analytical monitoring:

not required

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: 5 min ultrasound at room temperature
- Differential loading: no
- Controls: yes, freshly washed activated sludge without test item
- Evidence of undissolved material: yes, the test item was concentration related partially dispersed. Many test item particles swam on the surface of the water.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Laboratory culture: no
- Name and location of sewage treatment plant where inoculum was collected: Hildesheim, Germany
- Preparation of inoculum for exposure: 200 mL inoculum, 16 mL synthetic waste water, 284 mL demineralised water were mixed with respective test item concentrations (directly weighed)
- Pretreatment: The sludge was washed with autoclaved tap water and diluted corresponding to a dry weight of 3.0 ± 0.6 g/L.
- Initial biomass concentration: 3.02 g/L dry sludge

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Test temperature:

20.9 °C

pH:

6.89 (activated sludge)

Nominal and measured concentrations:

nominal: 1000, 1800, 3200, and 5800 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 500 mL Erlenmeyer flasks, DIN 12440
- size, headspace, fill volume: Volume = 500 mL; Volume study medium = 500 mL; Headspace = 0 mL
- Aeration: permanent
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2
- Sludge concentration: 3.02 g/L dry sludge
- Nutrients provided for bacteria: synthetic waste water according to OECD 209

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

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Temperature in the measuring cell: measured once during the measurement
- Room temperature [°C]: measured continuously

EFFECT PARAMETERS MEASURED:
- pH of the activated sludge: before testing
- oxygen depletion of the control, reference and test item: after 3 h incubation during at least 3 min

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
- Range finding study: yes
- Test concentrations: 10, 100, and 1000 mg/L
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

Copper (II) sulphate pentahydrate

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 5 800 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

- Effect concentrations exceeding solubility of substance in test medium: yes
- Respiration rate [mgO2/L*h] in control: 34.0, 32.4

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: EC50 (3 h) = 119 mg/L Copper (II) sulphate pentahydrate

Reported statistics and error estimates:

EC values were calculated with probit analysis using SigmaPlot rel. 6.0 (2000, SPSS Corporation).
Probit values were taken from Weber (1986).
Confidence intervals were calculated according to a standard procedure (Breitig & Tümpling 1982).

Table 1. Respiration rates and inhibition of respiration of test and reference item concentrations.

	Concentration [mg/L]	Respiration rate [mg O2/L*h]	Inhibition [%]
Test item	10000	---^1)	---^1)
	5800	32.4	2
	3200	30.4	8
	1800	30.8	7
	1000	30.4	8
Control 1	--	34.0	—
Control 2	--	32.4	—
Reference item	180	14.4	67
	100	26.0	41
	58	32.8	25
Control 1	--	42.8	—
Control 2	--	45.2	—

¹⁾Not measured because test item particles blocked up the measuring system

Table 2. Raw data and respiration rates of test and reference item concentrations.

	Concentration [mg/L]	Oxygen-concentration^*[mg O2/L]	Measured distance^**[mm/3 min]	Respiration rate [mg O2/L*h]
Test item	10000	---^1)	---^1)	---^1)
	5800	4.41	81	32.4
	3200	4.05	76	30.4
	1800	3.72	77	30.8
	1000	3.31	76	30.4
Control 1	--	6.09	85	34.0
Control 2	--	6.64	81	32.4
Reference item	180	8.00	36	14.4
	100	6.72	65	26.0
	58	5.58	82	32.8
Control 1	--	4.27	107	42.8
Control 2	--	3.99	113	45.2

¹⁾not measured because test item particles blocked up the measuring system

^*determined at the start of the 3 min measurement

^**5 mm are related to 0.1 mg O2/L

 

Validity criteria fulfilled:

yes

Conclusions:

EC50 (3 h) > 5800 mg/L (nominal concentration, OECD 209, activated sludge)

Description of key information

EC50 (3 h) > 5800 mg/L (nominal concentration, OECD 209, activated sludge)

Additional information

One GLP certified guideline study is available, which assessed the effects of the test substance on microorganisms from activated sludge according to the OECD guideline 209.

In a static test, a fresh sample of non-adapted activated sludge (3.02 g dw/L) from a local municipal STP was inoculated with four nominal concentrations of the test substance ranging from 1000 - 5800 mg/L. After a contact time of 3 h respiration rates were measured for at least 3 min. At test termination the inhibitory effects were determined and compared to the negative control.

No biologically significant inhibition of respiration was measured in the tested concentrations up to 5800 mg/L. Consequently, no EC values could be determined in the tested concentration range up to 5800 mg/L and the derived EC50 (3 h) was > 5800 mg/L.