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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:
22-08-2019 to 12-02-2020
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study performed under GLP. All relevant validity criteria were met.
Qualifier:
according to guideline
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Japan guidelines : "II: Method for Testing the Biodegradability of Chemical Substances by Microorganisms (301F)"
Version / remarks:
Stipulated in: No.20190619-2, Manufacturing Industries Bureau, Ministry of Economy, Trade and Industry; No. 1907011, Environmental Policy Bureau, Ministry of the Environment, Japan) and other citations within the full study report.
Deviations:
not specified
GLP compliance:
yes
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Activated sludge was obtained : from an aeration tank of a sewage treatment plant treating predominantly domestic sewage in Kurume-shi, Fukuoka, Japan. The sample was collected the day before test item exposure. The activated sludge of a (domestic) waste water treatment plant is recommended by test guideline.
- Storage conditions: See pretreatment field.
- Storage length: ca. 1 day from sampling to test initiation. The activated sludge was aerated at approximately 22°C until use for the test.
- Preparation of inoculum for exposure: See above.
- Pretreatment: Additive amount of the activated sludge into the test vessel was 4.23 mL on the basis of the concentration of suspended solid in the activated sludge which was determined by the following method: Method In accordance with Japanese Industrial Standards (JIS) K 0102:2016 : Section 14.1 (day before test initiation) : suspended solid concentration of activated sludge : 2130 mg/L , then diluted in test vessel to final test vessel suspended solid concentration : 30.0 mg/L).
- Concentration of sludge: The inoculum was added into the test suspensions, the procedure control, the inoculum blank and toxicity control to give a final concentration of 30.0 mg/L (dry weight).
- Water filtered: Not reported.
- Type and size of filter used, if any: Not reported.
Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Parameter followed for biodegradation estimation:
DOC removal
Parameter followed for biodegradation estimation:
other: Percentage biodegradation of test item by GC
Details on study design:
TEST CONDITIONS
- Composition of medium: Activated sludge was obtained : from an aeration tank of a sewage treatment plant treating predominantly domestic sewage in Kurume-shi, Fukuoka, Japan. The sample was collected the day before test item exposure. The activated sludge of a (domestic) waste water treatment plant is recommended by test guideline. Additive amount of the activated sludge into the test vessel was 4.23 mL on the basis of the concentration of suspended solid in the activated sludge which was determined by the following method: Method In accordance with Japanese Industrial Standards (JIS) K 0102:2016 : Section 14.1 (day before test initiation) : suspended solid concentration of activated sludge : 2130 mg/L , then diluted in test vessel to final test vessel suspended solid concentration : 30.0 mg/L). The mineral medium (3 L) was prepared at the same proportion as the following method: 10 mL of solution A and each 1 mL of solutions B, C, and D prescribed in the test methods in Section 5.
were made up to 1 L with purified water (The Japanese Pharmacopoeia, Takasugi Pharmaceutical), and then the pH of this solution was adjusted to 7.4.
- Additional substrate: Not applicable.
- Solubilising agent (type and concentration if used): Not applicable.
- Test temperature: 22 ±1 °C
- pH: Blank (water + test item): not reported ; Sludge + test item: 7.3 ; control blank: 7.3
- pH adjusted: No.
- CEC (meq/100 g): Not reported.
- Aeration of dilution water: Not reported.
- Suspended solids concentration: 30 mg SS/L.
- Continuous darkness: Yes.
- Other: Not applicable.

TEST SYSTEM
- Culturing apparatus: Glass vessel (ca. 300 mL).
- Number of culture flasks/concentration: Duplicate.
- Method used to create aerobic conditions: Not applicable.
- Method used to create anaerobic conditions: Not applicable.
- Measuring equipment: See previous sections.
- Test performed in closed vessels due to significant volatility of test substance: Not reported (although suspected)
- Test performed in open system: Not applicable.
- Details of trap for CO2 and volatile organics if used: Not applicable.
- Other: Not applicable.

SAMPLING
- Sampling frequency: BOD (oxygen consumption) was monitored continuously ; minimum end of incubation for DOC and/or GC for parent (if necessary) transformation products analysis.
- Sampling method: See above.
- Sterility check if applicable: Not applicable.
- Sample storage before analysis: Not applicable.
- Other: Not applicable.

CONTROL AND BLANK SYSTEM
- Inoculum blank: Yes.
- Abiotic sterile control: Not applicable.
- Toxicity control: Yes.
- Other: Reference item control (Sodium benzoate)

STATISTICAL METHODS: Not applicable.
Reference substance:
benzoic acid, sodium salt
Remarks:
100 mg/L
Test performance:
1. The repeatability validity criterion (not more than 20% difference between replicates) is fulfilled. Therefore, the test is considered valid (Actual: 2% in BOD and 1% in GC)
2. The BOD of the inoculated blank control was 29 mgO2/L and < 60 mgO2/L after 28 days
3. The pH at day 28 was in the range of 6.0 to 8.5 (actual 7.3 for test item vessels)
4. Sodium benzoate attained 87% degradation after 14 days thereby confirming the suitability of the inoculum and test conditions.
5. The percentage degradation of the toxicity control (sludge + test item + sodium benzoate) was 28% which had reached the pass level 25% of ThOD on 14 days.
All validity criteria were considered to be met.
Key result
Parameter:
% degradation (O2 consumption)
Remarks:
BOD ; mean
Value:
0
Sampling time:
28 d
Remarks on result:
other: actual mean = -6% (n=2 ; -5% and -7%) which is taken to indicate 0% biodegradation by BOD
Key result
Parameter:
% degradation (test mat. analysis)
Remarks:
GC ; mean
Value:
0
Sampling time:
28 d
Details on results:
The test item was degraded based on BOD: 0% at day 28 [actual mean = -6% (n=2 ; -5% and -7%) which is taken to indicate 0% biodegradation by BOD].
The test item was degraded based on GC: 0% at day 28.
Results with reference substance:
Sodium benzoate attained 87% degradation after 14 days thereby confirming the suitability of the inoculum and test conditions.
Validity criteria fulfilled:
yes
Interpretation of results:
not readily biodegradable
Conclusions:
The mean biodegradation in duplicate was 0 % at day 28 by BOD. The mean biodegradation by direct analysis by GC was 0% at day 28. Transformation and biodegradation of the test item did not occur under the test conditions of this study.
Executive summary:

The ready biodegradability test was carried out according to Japanese guidelines and OECD TG 301F guideline under GLP. The test item, at a concentration of 100 mg/L was exposed to mixed activated sewage sludge micro-organisms obtained from the activated sludge from an aeration tank of a sewage treatment plant treating predominantly domestic sewage in Kurume-shi, Fukuoka, Japan. The sample was collected the day before test item exposure. The activated sludge, was prepared in laboratory according to Japan guidelines. The test item was exposed in culture medium in sealed culture vessels in the dark at 22°C ± 1°C for 28 days. The sludge was diluted in the BOD bottles to 30 mg SS/L at exposure volume of 300 mL. The degradation of the test item was assessed by the measurement of oxygen consumption (BOD) on days 0 through 28 using a closed system oxygen consumption apparatus. DOC was determined quantitatively by TOC analyser. The test item degradation was determined directly quantitatively by Gas Chromatography (GC-FID) and/or DOC analysis. Control solutions with inoculum and the reference substance, sodium benzoate, together with blank controls were used for validation purposes. In the test inoculum blank the oxygen uptake was 29 mg O2/L. The pH value at the end of the test period 28 days did not exceed 7.3 in the test item system and the blank control system. The repeatability validity criterion of not more than 20% difference between replicates was fulfilled. Therefore, the test is considered valid. The test system met the validation criteria of the guideline. Sodium benzoate attained 87% degradation after 14 days thereby confirming the suitability of the inoculum and test conditions. The mean biodegradation for duplicate test flasks at 28 days for the test item was 0% (actual -5% and -7%) by BOD. The mean biodegradation by direct analysis by GC was 0% at day 28. No degradation products were detected within the chromatograms. Detected DOC corresponded to the test item dissolved in the test solutions. Under the conditions of the study, test item is not considered as readily biodegradable.

Description of key information

Biodegradation: not readily biodegradable ; mean biodegradation 0 % by BOD and GC (28 -days); transformation products were detected by GC, OECD TG 301F, 2020

Key value for chemical safety assessment

Biodegradation in water:
not biodegradable
Type of water:
freshwater

Additional information

Key study : OECD TG 301F, 2020 : The ready biodegradability test was carried out according to Japanese guidelines and OECD TG 301F guideline under GLP. The test item, at a concentration of 100 mg/L was exposed to mixed activated sewage sludge micro-organisms obtained from the activated sludge from an aeration tank of a sewage treatment plant treating predominantly domestic sewage in Kurume-shi, Fukuoka, Japan. The sample was collected the day before test item exposure. The activated sludge, was prepared in laboratory according to Japan guidelines. The test item was exposed in culture medium in sealed culture vessels in the dark at 22°C ± 1°C for 28 days. The sludge was diluted in the BOD bottles to 30 mg SS/L at exposure volume of 300 mL. The degradation of the test item was assessed by the measurement of oxygen consumption (BOD) on days 0 through 28 using a closed system oxygen consumption apparatus. DOC was determined quantitatively by TOC analyser. The test item degradation was determined directly quantitatively by Gas Chromatography (GC-FID) and/or DOC analysis. Control solutions with inoculum and the reference substance, sodium benzoate, together with blank controls were used for validation purposes. In the test inoculum blank the oxygen uptake was 29 mg O2/L. The pH value at the end of the test period 28 days did not exceed 7.3 in the test item system and the blank control system. The repeatability validity criterion of not more than 20% difference between replicates was fulfilled. Therefore, the test is considered valid. The test system met the validation criteria of the guideline. Sodium benzoate attained 87% degradation after 14 days thereby confirming the suitability of the inoculum and test conditions. The mean biodegradation for duplicate test flasks at 28 days for the test item was 0% (actual -5% and -7%) by BOD. The mean biodegradation by direct analysis by GC was 0% at day 28. No degradation products were detected within the chromatograms. Detected DOC corresponded to the test item dissolved in the test solutions. Under the conditions of the study, test item is not considered as readily biodegradable.