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Environmental fate & pathways

Biodegradation in water: screening tests

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Administrative data

biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
October 09 - November 07, 1996
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study has been performed according to OECD and EC guidelines and according to GLP principles.

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guidelineopen allclose all
according to guideline
other: Directive 92/69/EEC, C.4-C Carbon dioxide evolution test (Modified Sturm test)
according to guideline
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Details on test material:
- Name of test material (as cited in study report): Nikkol VC-IP
- Substance type: Colourless to light yellow transparent liquid
- Physical state: Liquid
- Stability under test conditions: stable
- Storage condition of test material: In refrigerator in the dark

Study design

Oxygen conditions:
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- The source of test organisms was activated sludge freshly obtained from a municipal sewage treatment plant: 'Waterschap de Aa', Schijndel,
the Netherlands.
- Before use, the sludge was allowed to settle for at least 30 minutes and the liquid decanted for use as inoculum at the amountof 10 ml/l of mineral
- Pretreatment: no special pretreatment
- Concentration of sludge: 10 ml supernatant/l of mineral medium.
- Initial cell/biomass concentration: not determined
- Water filtered: no

Duration of test (contact time):
29 d
Initial test substance concentrationopen allclose all
Initial conc.:
32.1 other: mg / 2 L
Based on:
test mat.
Initial conc.:
12 mg/L
Based on:
other: total organic carbon (TOC)
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
- Composition of medium: 1 litre mineral medium contains: 10 ml of solution (A), 1 ml of solutions (B) to (D)
and Milli-RO water.
Stock solutions of mineral components:
A) 8.50 g KH2PO4; 21.75 g K2HPO4; 67.20 g Na2HPO4.12H2O; 0.50 g NH4Cl; dissolved in Milli-Q water
and made up to 1 litre, pH 7.4 ± 0.2
B) 22.50 g MgSO4.7H2O dissolved in Milli-Q water and made up to 1 litre.
C) 36.40 g CaCl2.2H2O dissolved in Milli-Q water and made up to 1 litre.
D) 0.25 g FeCl3.6H2O dissolved in Milli-Q water and made up to 1 litre.
- Test temperature: between 20 and 21°C
- pH: Just before the start of the test: 7.6. On day 28: 7.6-7.9
- pH adjusted: no
- Aeration of dilution water: not before the test, the test is aerated continuously
- Suspended solids concentration: The concentration of suspended solids was 3.6 g/l in the concentrated
sludge (information obtained from the municipal sewage treatment plant).
- Continuous darkness: yes

- Culturing apparatus: Test vessels: 2 litre all-glass brown coloured bottles
- Number of culture flasks/concentration: Test suspension: containing test substance and inoculum (2
Inoculum blank: containing only inoculum (2 bottles)
Positive control: containing reference substance and inoculum (1 bottle)
Toxicity control: containing test substance, reference substance and inoculum (1 bottle).
- Method used to create aerobic conditions: A mixture of oxygen (21%) and nitrogen (79%) was passed
through a bottle, containing 0.5 - 1 litre 0.0125 M Ba(OH)2 solution to trap CO2 which might be present in
small amounts. The synthetic air was sparged through the scrubbing solutions at a rate of approximately 1-2
bubbles per second (ca. 30-100 ml/min).
- Test performed in open system: yes
- Details of trap for CO2 and volatile organics if used: The CO2 produced in each test bottle reacted with the
barium hydroxide in the gas scrubbing bottle and precipitated out as barium carbonate. The amount of CO2
produced was determined by titrating the remaining Ba(OH)2 with 0.05 M standardized HCl (1:20 dilution
from 1 M HCl (Titrisol® ampul), Merck, Darmstadt, Germany). Titrations were made every second or third day
during the first 10 days, and thereafter at least every fifth day until the 28th day. Each time the CO2-absorber
nearest to the test bottle was removed for titration; each of the remaining two absorbers was moved one
position in the direction of the test bottle. A new CO2-absorber was placed at the far end of the series.
Phenolphthalein (1% solution in ethanol, Merck, Darmstadt, Germany) was used as pH-indicator. On the 28th
day, the pH of the test suspensions was measured and 1 ml of concentrated HCl (37%, Merck, Darmstadt,
Germany) was added to each bottle. The bottles were aerated overnight to drive off CO2 present in the test
suspension. The final titration was made on day 29.

- Sampling frequency: titrations were made on days: 2, 5, 7, 9, 14, 19, 23, 27 and 29.
- Sampling method: titration of whole volume of CO2-absorber
- Inoculum blank: yes
- Abiotic sterile control: no
- Toxicity control: yes
Reference substance
Reference substance:
other: Sodium acetate

Results and discussion

% Degradation
% degradation (CO2 evolution)
ca. 6.1
Sampling time:
28 d
Details on results:
Points of degradation plot (test substance):
0 % degradation after 2 d
.5 % degradation after 5 d
2.8 % degradation after 7 d
2.8 % degradation after 9 d
2.9 % degradation after 14 d
3.2 % degradation after 19 d
3.2 % degradation after 23 d
4.2 % degradation after 27 d
5.2 % degradation after 29 d

BOD5 / COD results

Results with reference substance:
In the toxicity control more than 25% degradation occurred in 14 days (based on ThCO2). Therefore, the test substance was assumed to be not
Points of degradation plot (reference substance):
5.6 % degradation after 2 d
40.2 % degradation after 5 d
56.1 % degradation after 7 d
64.9 % degradation after 9 d
75.5 % degradation after 14 d
79.7 % degradation after 19 d
81.1 % degradation after 23 d
82.1 % degradation after 27 d
84.6 % degradation after 29 d

Applicant's summary and conclusion

Interpretation of results:
under test conditions no biodegradation observed
NIKKOL VC-IP was not readily biodegradable under the conditions of the modified Sturm test presently performed.