Fatty acids, coco, iso-Bu esters

Toxicity to microorganisms

Administrative data

Endpoint:

activated sludge respiration inhibition testing

Type of information:

migrated information: read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Study period:

01 Jul 2010

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

GLP - Guideline study, tested with the source substance ethyl linoleate (CAS No. 544-35-4). According to the ECHA guidance document “Practical guide 6: How to report read-across and categories (March 2010)”, the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Test guideline 3

Qualifier:

according to guideline

Guideline:

ISO 8192 (Water quality - Test for inhibition of oxygen consumption by activated sludge for carbonaceous and ammonium oxidation)

GLP compliance:

yes (incl. QA statement)

Remarks:

Food and Consumer Product Safety Authority, Den Haag, The Netherlands

Test material

Test material information

Constituent 1

Reference substance name:

Ethyl linoleate

EC Number:

208-868-4

EC Name:

Ethyl linoleate

Cas Number:

544-35-4

Molecular formula:

C20H36O2

IUPAC Name:

ethyl octadeca-9,12-dienoate

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

A development and validation of an analytical method for the test substance in ISO medium was performed
- Concentrations: 0.5 and 12.0 mg/L
- Sampling method: additional test, separate samples to respiration inhibition test
- Sample storage conditions before analysis: fresh and stored at room temperature for at least 6 days

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Since the test substance was hardly soluble in water, test concentrations were prepared separately in Milli-RO water (tap water purified by reverse osmosis). Two 1-L test bottles were filled with 250 mL test substance mixtures with initial loading rates of 200 mg/L. These mixtures were stirred in closed dark brown glass bottles for 25 hours. Subsequently, 16 mL synthetic sewage feed, 200 mL sludge and Milli-RO water up to 500 mL were added resulting in a loading rate of 100 mg/L.

Test organisms

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Source: Municipal sewage treatment plant: "Waterschap Aa en Maas', 's-Hertogenbosch, The Netherlands, receiving mainly domestic sewage
- Method of cultivation: Sludge was used one day after collection and kept aerated at test temperature until use
- Preparation of inoculum for exposure: Sludge was coarsely sieved, washed and diluted with ISO-medium. A small amount of sludge was weighed and dried overnight at ca. 105 °C to determine the amount of suspended solids
- Initial biomass concentration: 3.8 g suspended solids/L of sludge

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Test conditions

Test temperature:

18.3-18.9°C

pH:

8.0 - 8.3

Dissolved oxygen:

constant aeration

Nominal and measured concentrations:

0 and 100 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
- Test vessel: 1L test bottles (glass)
- Volume: 500 mL
- constant stirring and aeration with clean oil-free air via pipette
- No. of vessels per concentration (replicates): 2
- No. of controls: 2
- No. of vessels per reference substance (replicates): three concentrations (5, 12 and 30 mg/L)

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Milli-RO water
- Following the overnight stirring period, synthetic sewage feed (16 mL) and the activated sludge (200 mL) were added to the test substance mixture. Milli-RO water was added to provide a final volume of 500 mL.
- Composition of synthetic sewage feed: 16 g peptone, 11 g meat extract, 3 g urea, 0.7 g NaCl, 0.4 g CaCl2*2H2O, 0.2 g MgSO4*7H2O, 2.8 g K2HPO4; made up to 1 L dissolved in MilliQ water, pH was 7.5 ± 0.5

EFFECT PARAMETERS MEASURED: inhibition of the respiration rate
- after 3 hour contact time, a well mixed sample of the content was poured into a 300 mL oxygen bottle (glass) and the flask was sealed with an oxygen electrode connected to a recorder (flatbed recorder SE 102), forcing the air out of the vessel. Oxygen consumption was measured and recorded for approx. 10 minutes. During measurement, no aeration took place, but there was constant stirring via magnetic stirrer.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Results and discussion

Effect concentrations

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

- The difference between controls was within 15% of each other
- Nominal analytical concentrations of 0.5 and 10.0 mg/L were analysed as 0.424, 0.393, 0.411, 0.362 and 0.406 mg/L and as 12.0, 11.6, 9.91, 10.2 and 10.3 mg/L, respectively

Results with reference substance (positive control):

- The EC50 of 3,5-dichlorophenol was in the accepted range of 5-30 mg/L (9.3 mg/L)
- Regression line y = 51.34 x + 0.3, r2 = 0.9950

Any other information on results incl. tables

Oxygen concentration and oxygen consumption:

Flask	Conc. of test substance (mg/L)	Oxygen conc. at the start (mg O2/L)	Oxygen consumption (mg O2/L/h)	% Inhibition respiration rate
control 1	0	7.6	41	-
control 2	0	8.0	42	-
T1	100	5.3	45	-8*
T2	100	6.9	42	-*1

* A negative value indicates stimulation of the sludge. This value is considered as not significant