Linseed oil, polymd.

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

18 September 2013 - 20 November 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

in accordance with GLP

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)

Deviations:

yes

Remarks:

(Prolongation up to 60 days, ammonium chloride omitted from medium to prevent oxygen consumption due to nitrification (no nitrogen limitation as shown by biodegradation of reference compound), inoculum activated sludge instead of effluent/surface water)

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: Secondary activated sludge (12-09-13) from the STP Nieuwgraaf in Duiven, The Netherlands - treating predominatntly domestic wastewater.
- Storage conditions: activated sludge was preconditioned to reduce the endogenous respiration rates: 400 mg dry Weight (DW)/L of activated sludge was aerated for one week. Sludge was diluted in the biological oxygen demand (BOD) bottles (van Ginkel and Stroo, 1992).

Duration of test (contact time):

60 d

Initial conc.:

2 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium:
Nutrients per liter of deionized water:
8.5 mg KH2PO4,
21.75 mg K2HPO4,
33.4 mg Na2HPO4•2H2O,
22.5 mg MgSO4•7H2O,
27.5 mg CaCl2,
0.25 mg FeCl3•6H2O.
Ammonium chloride was omitted to prevent nitrification.

- Solubilising agent: Dichloromethane (Sigma-Aldrich)
Sodium acetate was added to the bottles using an aqueous stock solution of 1.0 g/L. Linseed standoil was added to the bottles using a stock of 1.0 g/L in DCM.
For the Closed Bottle test, the test substance in DCM (0.6 mL) was directly added to the bottles. The bottles were placed on a roller bank in a ventilated hood for a few hours to allow the solvent to evaporate and to obtain an even distribution of the test substance on the walls of the bottles.
- Test temperature: 22°C -24°C
- pH: at day 0: 7.2
at day 28: 7.3 (both controls) and 7.2 (test)
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 0.30 L BOD (biological oxygen demand) bottles with glass stoppers
- Number of culture flasks/concentration: 10 bottles containing only inoculum; 6 bottles containing inoculum and sodium acetate; 10 bottles containing inoculum and test substance; and 10 bottles treated with DCM containing inoculum.
- Measuring equipment: electronical measurement of the oxygen concentration using an oxygen electrode (WTW TrioXmatic EO 200) and meter (WTW OXI 530) (Retsch, Ochten, The Netherlands); pH was measured using a Eutech Cyberscan pH11 pH meter (Eutech Instruments, Nijkerk, The Netherlands)

SAMPLING
- Sampling frequency: day 0, 7, 14, 21 and 28
- Sampling method: two duplicate bottles of all series were withdrawn for dissolved oxygen analyses on the respective days; One extenstion of the Closed bottle test by measuring the course of the oxygen decrease in the bottles of day 28 using a special funnel. An oxygen electrode was inserted in the BOD bottle to measure the oxygen concentration. Medium dissipated by the electrode flowed back into the BOD bottle after withdrawal of the oxgen electrode from the funnel (van ginkel and Stroo 1992). This method allowed measurement at day 42 and 60.

CONTROL AND BLANK SYSTEM
- Inoculum blank: inoculum alone and inoculum with DCM

Key result

Parameter:

% degradation (O2 consumption)

Value:

33

Sampling time:

28 d

Key result

Parameter:

% degradation (O2 consumption)

Value:

41

Sampling time:

60 d

Remarks on result:

other: prolonged Closed Bottle test

Details on results:

See table under 'Any other information'
The calculated theoretical oxygen demand (ThOD) of linseed standoil is 2.9 mg/mg and the ThOD of sodium acetate is 0.8 mg/mg.
The inhibition of the degradation of sodium acetate by the test substance in the Closed Bottle test was not determined due to irrelevent possible toxicity of linseed standoil to microorganisms degrading acetate. Inhibition of the endogenous respiration of the inoculum by the test substance at day 7 was not detected. Thus, no inhibition of the biodegradation expected.

Results with reference substance:

The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 81.

Oxygen consumption (mg/L) and biodegradation (%) of linseed standoil (BOD/ThOD) and sodium acetate (BOD/ThOD) in the Closed Bottle test.

Time (days)	Oxygen consumption (mg/L)		Biodegradation (%)
	Test substance	Acetate	Test substance	Acetate
0	0.0	0.0	0	0
7	0.9	4.1	19	76
14	1.3	4.4	22	81
21	1.7		29
28	1.9		33
42	2.0		34
60	2.4		41

Validity criteria fulfilled:

yes

Remarks:

Difference of the replicate values at day 28 < 20%; Biodegradation percentage of the reference compound (sodium acetate) was 81 at day 14; Oxygen concentration > 0.5 mg/L in all bottles during the test period.

Interpretation of results:

inherently biodegradable

Conclusions:

Linseed standoil is biodegraded by 33% at day 28 and 41% at day 60 (prolonged Closed Bottle test). Linseed standoil is therefore not readily biodegradable, but can be classified as inherently biodegradable.

Executive summary:

A ready biodegradability test in an aerobic aqueous medium was conducted to assess the biotic degradation of linseed standoil. The used Closed Bottle test was performed according to slightly modified OECD, EU and ISO Test Guidelines and in compliance with the OECD principles of Good Laboratory Practice.

Linseed standoil is considered non-inhibitory to the inoculum as it did not cause a reduction in the endogenous respiration. The Linseed stand was degraded by 33% on day 28 and 41% on day 60 in the Closed Bottle test. Thus the test substances revealed to not be readily biodegradable, but can be classified as inherently biodegradable.

The test fullfils the validity criteria by an endogenous respiration of 0.9 mg/L and by the total mineralization of the reference compound, sodium acetate. Sodium acetate was degraded by 81% of its theoretical oxygen demand after 14 days. Also, all bottles contained an oxygen concentration of >0.5 mg/L during the entire test period.

Description of key information

Not readily biodegradable: 33% mineralisation (oxygen consumption) in 28 days (OECD TG 301D, GLP study, Rel. 1). Prolongation of the study showed 41% mineralisation after 60 days. The test substance is considered to be inherently biodegradable.  

Key value for chemical safety assessment

Biodegradation in water:

inherently biodegradable, not fulfilling specific criteria

Additional information

The degradation of Linseed oil, polymerized was tested in different tests for screening ready biodegradability. Based on the results, it is concluded that Linseed oil, polymerized is not readily biodegradable; however, the test substance is inherently biodegradable, not fulfilling specific criteria. A study was designed to performed the bioavailability of the test substance using agitation, a solvent or dispersants. The table summarises the efforts and the results of all studies.

	Remarks	% Degradation	Referentie
OECD 301D Closed Bottle test	Inoculum: activated sludge	28d: 33% of ThOD 60d: 41% of ThOD	Van Ginkel 2013
OECD 301D Closed Bottle test	Inoculum: activated sludge OR river water   Dispersants or solvent: Synperonic, Genapol, Tween 80/85, DCM	28d: 15 – 36% of ThOD 56d: 21 – 45% of ThOD	Van Ginkel & Geerts 2014
OECD 301B CO2-evolution test	Inoculum: activated sludge	28d: 13-16% of ThOD	Desmares 2010
Repetitive Die Away test	Inoculum: activated sludge   Dispersants: Genapol PF-40 Nonylphenol.10EO.5PO 3 more additions after the first dose	14d: 20% of ThOD   21d, 28 d: 14% of ThOD 42d: 19% of ThOD	Balk & Meuwsen 1988
OECD302A Semi-Continuous Activated sludge test	Inoculum: activated sludge   Dispersant: Tween 80	31d: 70 – 74% carbon removal 70d: 58 – 65% carbon removal	Van Ginkel &Geerts 2014

(Remark: All ready biodegradation tests are conducted in the presence of oxygen)