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Environmental fate & pathways

Biodegradation in water: screening tests

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Reference
Endpoint:
biodegradation in water: inherent biodegradability
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
From June 13th to July 11th, 2006
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
The usage of information on Direct Blue 199_Na, which has the same main component and with a different counter ion, can be considered as suitable and appropriated because the difference in salification is expected to not influence the characteristics related to the specific end-point.
The impurity profile does not impact on the read across proposed. Details on the approach followed are included in the document attached to the IUCLID section 13.
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
OECD Guideline 302 B (Inherent biodegradability: Zahn-Wellens/EMPA Test)
Version / remarks:
adopted on 17 July 1992
Principles of method if other than guideline:
The study was conducted in accordance with ISO/IEC 17025
GLP compliance:
no
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: municipal biological waste water treatment plant, ARA Werdhölzli, CH-8048 Zürich.
- Pre-conditioning: not pre-conditioned.
- Adaptation: the activated sludge was used immediately after sampling from the treatment plant without adaptation.
- Preparation of inoculum for exposure: prior to the test, the sludge was washed twice with tap water.
Duration of test (contact time):
28 d
Initial conc.:
ca. 50.6 mg/L
Based on:
DOC
Initial conc.:
ca. 146 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
DOC removal
Details on study design:
TEST CONDITIONS
- Composition of medium: aerobic mineral salts medium prepared with double distilled water (conductivity: < 1.5 µS/cm; DOC: < 0.3 mg/l).
Stock solution I, 10 ml/l (pH 7.4): Ammonium chloride 0.50 g, Di-sodium hydrogenphosphate 33.40 g, Potassium dihydrogenphosphate 8.50 g, Di-potassium monohydrogenphosphate 21.75 g
Stock solution Il, 1 ml/l: Calcium chloride, anhydrous 27.50 g
Stock solution III, 1 ml/l: Magnesium sulfate 22.50 g
Stock solution IV, 1 ml/l: Iron (III) chloride 0.25 g
- Test temperature: 22 ± 0.5 °C
- Additional substrate: none, test substance or procedure control as sole organic carbon source.
- Continuous darkness: yes.
- Oxyge: it was assured that during the test the oxygen concentration was greater than 6 mg/I.
- pH: the pH-value was checked periodically and adjusted to pH 6.5-8.0 with NaOH or HCI, if necessary.

TEST SYSTEM
- Inoculum: 0.2 g/l dry metter of activated sludge in final mixture.
- Test unit: 1200 ml closed glass bottle containing a total volume of test solution of 800 ml; aerated with CO2-free air and fitted to gas-absorption bottles containing 125 ml of 0.13 M KOH.
- Test suspension: two replicates containing activated sludge, test medium and the test substance.
- Stirring: the test vessels were stirred (100 r.p.m.)
- Aeration: the test vessels were areated with synthetic C02-free air for the test period.

SAMPLING
- Sampling: the elimination of the test material was followed by DOC determinations at regular intervals. First samples were analyzed at the beginning of and 3 h after starting the test.
- Air control: the air leaving the individual vessels was passed through gas-absorption bottles filled with NaOH. The trapped CO2 was determined as inorganic carbon (IC).

CONTROL AND BLANK SYSTEM
- Inoculum blank: two replicates containing activated sludge and test medium
- Reference compound: one replicate containing activated sludge, the test medium and the diethylene glycol as ready biodegradable reference compound.
Reference substance:
diethylene glycol
Remarks:
47.6 mg/l as DOC
Parameter:
% degradation (DOC removal)
Value:
5
Sampling time:
28 d
Parameter:
other: % degradation (ThCO2)
Value:
3
Sampling time:
28 d
Details on results:
No biodegradation of the test item based on the CO2 evolution and calculated as percentage ThCO was observed.

No significant elimination of the test substance due to adsorption to the activated sludge, on the glass surface or other physico-chemical processes was found as determined by means of DOC measurements 3 h after the start of the test.
Results with reference substance:
The positive control, diethyleneglycol, showed 99 % biodegradation after 14 days of incubation thus confirming suitability of inoculum and test conditions.

DOC concentrations of test suspension, inoculum blank and procedure control and calculation of degradation data.

Inoculum blank (B)* Procedure control with diethylene glycol (R) Test suspension with test material (T)*
Time DOC DOC DOC net. Degradation DOC DOC net. Degradation
(days) (mg/I) (mg/I) (mg/I) (%) (mg/I) (mg/I) (%)
0 0.8 48.7 47.9 - 49.9 49.0 -
0.125 0.6 47.7 47.1 1 50.2 49.6 0
1 0.9 46.6 45.7 4 49.9 49.1 1
3 1.1 37.3 36.2 24 49.0 47.9 3
7 1.4 1.9 0.5 99 49.7 48.4 3
10 2.3 3.3 1.0 98 50.0 47.7 4
14 1.4 1.9 0.5 99 49.5 48.1 3
17 1.4 1.8 0.4 99 49.4 48.0 3
21 1.5 2.0 0.5 99 49.2 47.7 4
24 1.5 1.7 0.2 100 48.5 46.9 5
27 1.5 1.8 0.3 99 48.5 46.9 5
28 1.8 1.9 0.1 100 49.1 47.3 5

*Mean of two replicates

IC concentration, calculated from the concentrations in the gas absorption bottles, of test suspension, inoculum blank and procedure control and corresponding degradation data

Time Test suspension (IC) Procedure control with diethylene glycol IC Inoculum blank IC Biodegradation test suspension Biodegradation procedure control
(days) (mg/I) (mg/I) (mg/I) (% ThCO2) (% ThCO2)
0 nd nd nd - -
7 23.1 58.9 22.7 1 76
14 32.4 76.6 31.9 1 94
21 39.5 82.0 38.3 2 92
28 42.7 86.4 41.4 3 95
Validity criteria fulfilled:
yes
Remarks:
reference compound reached more than 7 % of DOC removal within 14 days
Interpretation of results:
not inherently biodegradable
Conclusions:
DOC removal (28 days): 5 %
Executive summary:

The biodegradability of the test compound exposed to activated sludge of a municipal sewage treatment plant was investigated under aerobic static conditions. The study was conducted according to the OECD guideline 302B. No significant elimination of the test substance due to adsorption to the activated sludge, on the glass surface or other physico-chemical processes was found as determined by means of DOC measurements 3 h after the start of the test. The positive control, diethyleneglycol, showed 99 % biodegradation after 14 days of incubation thus confirming suitability of inoculum and test conditions.

Based on the data of the individuaL DOC determinations no biodegradability of the test compound was observed after 28 days. Furthermore, no biodegradation of the test substance based on CO2 evolution and calculated as % ThCO2 was observed, confirming that no biodegradation occurred.

Conclusion

Based on these results, the test substance is not inherently biodegradable under the conditions, since less than 20 % degradation was attained after 28 days of contact time.

Description of key information

neither readily nor inherently biodegradable

Key value for chemical safety assessment

Biodegradation in water:
under test conditions no biodegradation observed

Additional information

The biodegradability potential of the Direct Blue 199 has been investigated using the available information on the Direct Blue 199_Na. The usage of information on Direct Blue 199_Na, which has the same main component and with a different counter ion, can be considered as suitable and appropriated because the difference in salification is expected to not influence the characteristics related to the specific end-point. The impurity profile does not impact on the read across proposed. Details on the approach followed are included in the document attached to the IUCLID section 13.

Direct Blue 199_Na was exposed to activated sludge of a municipal sewage treatment plant was investigated under aerobic static conditions. The study was conducted according to the OECD guideline 302B. No significant elimination of the test substance due to adsorption to the activated sludge, on the glass surface or other physico-chemical processes was found as determined by means of DOC measurements 3 h after the start of the test. The positive control, diethyleneglycol, showed 99 % biodegradation after 14 days of incubation thus confirming suitability of inoculum and test conditions. Based on the data of the individual DOC determinations no biodegradability of the test compound was observed after 28 days. Furthermore, no biodegradation of the test substance based on CO2evolution and calculated as % ThCO2was observed, confirming that no biodegradation occurred.

Since less than 20 % of degradation was attained after 28 days of contact time, Direct Blue 199_Na is considered as not inherently biodegradable.

The ration BOD5:COD of Direct Blue 199_Na resulted lower than 0.5 in all of the test available, confirming that Direct Blue 199_Na is not readily biodegradable.