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Environmental fate & pathways

Biodegradation in water: screening tests

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Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
March 25 - April 29, 1993
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
minor deviations from standard test guidelines and minor methodological deficiencies, which do not affect the quality of the relevant result.
Qualifier:
according to guideline
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Version / remarks:
A minor deviation of the test procedures described in the guidelines was introduced: instead of an effluent/extract/mixture, activated sludge was used as an inoculum. The activated sludge was preconditioned to reduce the endogenous respiration rates. To this end, the sludge (200 mg Dry Weight (DW/l) was aerated for one week. The sludge was diluted to a concentration of 2 mg DW/l in the BOD bottles. This method was described in a proposal for harmonizing ready biodegradability test protocols [J. Blok, A. de Morsier, P. Gerike, L. Reynolds and H. Wellens: 'Harmonisation of ready biodegradability tests'. Chemosphere 14 (1985) 1805-1820)].
Deviations:
yes
Remarks:
please refer to 'Version/remarks'
Qualifier:
according to guideline
Guideline:
EU Method C.6 (Degradation: Chemical Oxygen Demand)
Version / remarks:
A minor deviation of the test procedures described in the guidelines was introduced: instead of an effluent/extract/mixture, activated sludge was used as an inoculum. The activated sludge was preconditioned to reduce the endogenous respiration rates. To this end, the sludge (200 mg Dry Weight (DW/l) was aerated for one week. The sludge was diluted to a concentration of 2 mg DW/l in the BOD bottles. This method was described in a proposal for harmonizing ready biodegradability test protocols [J. Blok, A. de Morsier, P. Gerike, L. Reynolds and H. Wellens: 'Harmonisation of ready biodegradability tests'. Chemosphere 14 (1985) 1805-1820)].
Deviations:
yes
Remarks:
please refer to 'Version/remarks'
Qualifier:
according to guideline
Guideline:
other: ISO/TC 147/SC 5WG 4N152; German proposal for an ISO-standard. Water quality evaluation in an aqueous medium of the 'ultimate' aerobic biodegradability of organic compounds. Method by analysis of biochemical oxygen demand (closed bottle test).
Version / remarks:
A minor deviation of the test procedures described in the guidelines was introduced: instead of an effluent/extract/mixture, activated sludge was used as an inoculum. The activated sludge was preconditioned to reduce the endogenous respiration rates. To this end, the sludge (200 mg Dry Weight (DW/l) was aerated for one week. The sludge was diluted to a concentration of 2 mg DW/l in the BOD bottles. This method was described in a proposal for harmonizing ready biodegradability test protocols [J. Blok, A. de Morsier, P. Gerike, L. Reynolds and H. Wellens: 'Harmonisation of ready biodegradability tests'. Chemosphere 14 (1985) 1805-1820)].
Deviations:
yes
Remarks:
please refer to 'Version/remarks'
GLP compliance:
yes (incl. QA statement)
Remarks:
Akzo Research Laboratories Arnhem
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, adapted
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): WWTP Nieuwgraaf in Duiven, Netherlands; the WWTP Nieuwgraaf is an activated sludge plant treating predominantly domestic waste water.
- Laboratory culture: not applicable
- Method of cultivation: no data
- Storage conditions: no data
- Storage length: no data
- Preparation of inoculum for exposure: instead of an effluent/extract/mixture, activated sludge was used as an inoculum. The activated sludge was preconditioned to reduce the endogenous respiration rates.
- Pretreatment: the sludge (200 mg Dry Weight (DWJ/l) was aerated for one week
- Concentration of sludge: the sludge was diluted to a concentration of 2 mg DW/l in the BOD bottles.
- Water filtered: no
Duration of test (contact time):
28 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: 8.5 mg KH2PO4 21.75 mg K2HPO4 33.3 mg Na2HP04»2H20, 22.5 mg MgSO4x7H20, 27.5 mg CaCl2, 0.25 mg FeCl3x6H20; ammonium chloride was omitted from the medium to prevent nitrification.
- Additional substrate: no
- Solubilising agent (type and concentration if used): no
- Test temperature: 21°C ± 1°C
- pH: The pH of the media was 6.9 at the start of the test. The pH of the medium at day 28 is 6.4 to 6.5.
- pH adjusted: no
- CEC (meq/100 g): no data
- Aeration of dilution water: 24h
- Suspended solids concentration: 2 mg DW/l (inoculum)
- Continuous darkness: yes


TEST SYSTEM
- Culturing apparatus: BOD flasks
- Number of culture flasks/concentration: 10 bottles containing only inoculum, 10 bottles containing test substance and inoculum, and 6 bottles
containing sodium acetate and inoculum.
- Method used to create aerobic conditions: aeration of test water
- Measuring equipment: for determination of dissolved oxygen concentration oxygen electrode (WTW Trioxmatic EO 200) and meter (WTW OXI 530);
the pH was measured using a pH meter (Consort P207); the temperature was measured and recorded with a Control One (IBT, Rotterdam,
Netherlands).
- Test performed in closed vessels: yes


SAMPLING
- Sampling frequency: day 7, 14,21, and 28
- Sampling method: two duplicate bottles of all series were withdrawn for analyses of the dissolved oxygen concentration
- Sample storage before analysis: no data


CONTROL AND BLANK SYSTEM
- Inoculum blank: yes
- Abiotic sterile control: not measured
- Toxicity control: not measured


STATISTICAL METHODS: none
Reference substance:
acetic acid, sodium salt
Preliminary study:
not performed
Test performance:
The test was performed in 250 to 300 ml BOD (biological oxygen demand) bottles with glass stoppers. Use was made of 10 bottles containing only inoculum, 10 bottles containing test substance and inoculum, and 6 bottles containing sodium acetate and inoculum. The concentrations of the test compound and sodium acetate in the bottles were 2.0 and 6.7 mg/l, respectively. The inoculum was diluted to 2 mg DW/l in the closed bottles. Each of the prepared solutions was dispensed into the respective group of BOD bottles so that all bottles were completely filled without air bubbles. The zero time bottles were immediately analysed for dissolved oxygen using an oxygen electrode. The remaining bottles were closed and incubated at 21°C ± 1°C in the dark. Two duplicate bottles of all series were withdrawn for analyses of the dissolved oxygen concentration at day 7, 14,21, and 28.
Parameter:
% degradation (O2 consumption)
Value:
86
Sampling time:
28 d
Details on results:
The validity of the test is demonstrated by an endogenous respiration of 1.1 mg/l at day 28 (Table I). Furthermore, the differences of the replicate values of the control at day 28 were less than 20%. The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 81. Finally, the validity of the test is shown by oxygen concentrations >0.5 mg/l in all bottles during the test period.
Results with reference substance:
The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 81.
Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The test substance is biodegraded 86% at day 28 in the closed bottle test and should therefore be classified as readily biodegradable.
Executive summary:

The ready biodegradability was determined in the closed bottle test performed according to slightly modified OECD (301D), EEC (C.6) and ISO Test Guidelines.

The test substance caused no reduction in the endogenous respiration. Therefore, Nouryset 200 is considered to be non-inhibitory to the inoculum.

Nouryset 200 was biodegraded 86% at day 28 in the closed bottle test. Hence this compound should be classified as readily biodegradable.

The test is valid as shown by an endogenous respiration of 1.1 mg/1 and by the total mineralization of the reference compound, sodium acetate. Sodium acetate was degraded 81% of its theoretical oxygen demand after 14 days. Finally, the most important criterion was met by oxygen concentrations >0.5 mg/l in all bottles during the test period.

Description of key information

readily biodegradable: 86 % in 28 days (OECD 301D) and 73.2 %  in 28 days (OECD 301D), respectively

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable
Type of water:
freshwater

Additional information

This endpoint is filled from data from two adequate ready biodegradability closed bottle tests (van Cinkel, 1993; unknown, 1994). Results of these two OECD guideline studies (OECD 301D) indicate that 'diallyl 2,2'-oxydiethyl dicarbonate' is readily biodegradable. The biodegradability of the test material on day 28 was 86 % and 73.2 %, respectively.