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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:
Experimental starting date: 16 August 2006. Experimental completion date: 20 August 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Deviations:
yes
Remarks:
instead of an effluent/extract/mixture, activated sludge was used as an inoculum
Qualifier:
according to
Guideline:
ISO 10707 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)
Deviations:
yes
Remarks:
instead of an effluent/extract/mixture, activated sludge was used as an inoculum
Qualifier:
according to
Guideline:
EU Method C.6 (Degradation: Chemical Oxygen Demand)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
chemical name: Alkylbenzenesulphonate
trade name: Witconate P-1460
purity: 88.8%
batch/lot: No. 2140-5901
appearance: Paste
stability: Not relevant
solubility in water: Suspension at 1.0 g/L
storage: At ambient temperature in the dark

Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
Secondary activated sludge was obtained from the WWTP Nieuwgraaf in Duiven, The Netherlands
(16-08-2006). The WWTP Nieuwgraaf is an activated sludge plant treating
predominantly domestic waste water. 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, 400 mg Dry Weight (DW)/L of activated sludge was aerated
for one week. The sludge was diluted to a concentration of 2 mg DW/L in the BOD bottles
(van Ginkel and Stroo 1992).
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 procedures
The Closed Bottle test was performed according to the study plan. The study plan was
developed from ISO Test Guidelines (1994). 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 analyzed for dissolved oxygen using an
oxygen electrode. The remaining bottles were closed and incubated 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.

Analyses
The dissolved oxygen concentrations were determined electrochemically using an oxygen
electrode (WTW Trioxmatic EO 200) and meter (WTW OXI 530) (Retsch, Ochten, The
Netherlands). The pH was measured using a Knick 765 calimatic pH meter (Elektronische
Messgerate GmbH, Berlin, Germany). The temperature was measured and recorded with a
thermo couple connected to a data logger.
The dry weight (DW) of the inoculum was determined by filtrating 50 ml of the activated
sludge over a preweighed 12 μm Schleicher and Schull filter. This filter was dried for
1.5 hours at 104°C and weighed after cooling. DW was calculated by subtracting the
weighed filters and by dividing this difference by the filtered volume.

Deionized water
Deionized water containing no more than 0.01 mg/L Cu (Sterlab certified; non-GLP
analysis) was prepared in a water purification system.

Test bottles
The test was performed in 250 to 300 ml BOD (biological oxygen demand) bottles with
glass stoppers.

Nutrient solution and stock solutions
The nutrient medium of the Closed Bottle test contained per liter of deionized water:
8.5 mg KH2PO4, 21.75 mg K2HPO4, 33.3 mg Na2HPO4·2H2O, 22.5 mg MgSO4·7H2O,
27 .5 mg CaCl2, 0.25 mg FeCl3·6H2O. Ammonium chloride was omitted from the medium to
prevent nitrification. Due to this omission the pH of the medium decreased slightly. The
decrease of the pH does not effect the biodegradation in the Closed Bottle test. The test
compound and sodium acetate were added to the bottles using a stock suspension and
stock solutions of 1.0 g/L, respectively.
Reference substance:
acetic acid, sodium salt
Key result
Parameter:
% degradation (O2 consumption)
Value:
83
Sampling time:
28 d
Details on results:
THOD
The calculated theoretical oxygen demand of Witconate P-1460 is 2.7 mg/mg. This
theoretical oxygen demand is calculated by assuming formation of carbon dioxide,
sulphate, nitrate and water. The theoretical oxygen demand of sodium acetate is
0.8 mg/mg.

Toxicity
Inhibition of the degradation of a well-degradable compound, e.g. sodium acetate by the
test compound in the Closed Bottle test was not determined because possible toxicity of
Witconate P-1460 to microorganisms degrading acetate is not relevant. Inhibition of the
endogenous respiration of the inoculum by the test substance was not detected. Therefore,
no inhibition of the biodegradation due to the "high" initial concentration of the test
compound is expected.

Test conditions
The pH of the media was 7 .0 at the start of the test. The pH of the medium at day 28 was
6.8. Temperatures ranged from 19 to 21 °C.

Validity of the test
The validity of the test is demonstrated by an endogenous respiration of 0.6 mg/Lat day
28. Furthermore, the differences of the replicate values at day 28 were less than
20%. The biodegradation percentage of the reference compound, sodium acetate, at day
14 was 85. Finally, the validity of the test is shown by oxygen concentrations >0.5 mg/Lin
all bottles during the test period.
Key result
Parameter:
ThOD
Value:
2.7 other: mg/mg
Results with reference substance:
The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 85%.

Oxygen consumption (mg/L) and the percentages biodegradation of Witconate P-1460 (BOD/ThOD) and sodium acetate (BOD/ThOD) in the Closed Bottle test.

Time (Days) Oxygen consumption (mg/L) Biodegradation (%)
Test Acetate Test Acetate
0 0 0 0 0
7 2.3 4.1 43 76
14 3.9 4.6 72 85
21 4.3 80
28 4.5 83
Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
Witconate P-1460 is biodegraded 83% at day 28 in the Closed Bottle test and should therefore be classified as readily biodegradable.
Executive summary:

In order to assess the biotic degradation, a ready biodegradability test was performed

which allows the biodegradability to be measured in an aerobic aqueous medium. The

ready biodegradability was determined in the Closed Bottle test performed according to

slightly modified OECD 301, EU C.6 and ISO 10707 Test Guidelines, and in compliance with the OECD

principles of Good Laboratory Practice.

Witconate P-1460 did not cause a reduction in the endogenous respiration. The test

substance is therefore considered to be non-inhibitory to the inoculum.

Witconate P-1460 was biodegraded 83% 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 0.6 mg/L and by the total

mineralization of the reference compound, sodium acetate. Sodium acetate was degraded

85% of its theoretical oxygen demand after 14 days. Finally, the most important criterion

was met by oxygen concentrations >0.5 mg/Lin all bottles during the test period.

Description of key information

In order to assess the biotic degradation, a ready biodegradability test was performed which allows the biodegradability to be measured in an aerobic aqueous medium. The ready biodegradability was determined in the Closed Bottle test performed according to slightly modified OECD 301, EU C.6 and ISO 10707 Test Guidelines, and in compliance with the OECD principles of Good Laboratory Practice.

Benzenesulfonic acid, mono-C10-13-alkyl derivs., compds. with N1,N1-dimethyl-1,3-propanediamine did not cause a reduction in the endogenous respiration. The test substance is therefore considered to be non-inhibitory to the inoculum.

Benzenesulfonic acid, mono-C10-13-alkyl derivs., compds. with N1,N1-dimethyl-1,3-propanediamine was biodegraded 83% 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 0.6 mg/L and by the total mineralization of the reference compound, sodium acetate. Sodium acetate was degraded 85% 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.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

[Type of water: freshwater]