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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:
2010-11-15 to 2010-12-13
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
other: Biodegradability Test of chemical Substances by Microorganisms (Yakushokuhatsu No. 1121003, Heisei 15.11.17 Seikyoku No.3, Kanpokihatsu No.031121004, November 21, 2003, the latest revision, July 4, 2008)
Deviations:
no
GLP compliance:
yes
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: Chemicals Evaluation and Research Institute, Japan
- Concentration of sludge: 30 mg/L
- Storage length: 6 days- date of receipt November 9, 2010
- Mixed liquor suspended solid (MLSS): 3800 mg/L BOD measurements 3600 mg/L recovery test
Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
DOC removal
Remarks:
dissolved organic carbon
Parameter followed for biodegradation estimation:
other: BOD biochemical oxygen demand
Details on study design:
TEST CONDITIONS
- Composition of medium:
Basal medium: Purified water (grade 4) aqueous solution containing 0.3 % each of liquid A, B, C, D adjusted pH to 7.0 ± 0.04 with 1 N H3PO4 or 1 N NaOH
Solution A: Aqueous solution containing 2.175 % K2HPO4, 0.85 % KH2PO4, 4.46 % Na2HPO4*12 H2O and 0.17 % NH4Cl
Solution B: Aqueous solution of 2.25 % MgSO4*7H2O
Solution C: Aqueous solution of 2.75 % CaCl2
Solution D: Aqueous solution of 0.025 % FeCl3*6H2O
-Test volume: 300 mL
- Stirring: Continuous stirring with magnetic stirrer
- Solubilising agent: Purified water grade 4
- Test temperature: 25 ± 1 °C
- pH: 7.0 ± 0.5 pH measurement at the start and end of the exposure period;before the exposure inoculum blank, purified water; after the exposure all test bottles
- pH adjusted: no
- Suspended solids concentrations:test substance (bottles 3-6) 100mg/L; aniline(bottle) 100 mg/L activated sludge (bottles 1-5) 30 mg/L

TEST SYSTEM
- Number of culture flasks/concentration: 3 duplicates
Bottle 1 (activity control): aniline+sludge+basal medium
Bottle 2 (inoculum blank): sludge+basal medium
Bottles 3,4,5 (test suspensions-1, 2, 3): test substance(30 mg)+ sludge + basal medium
Bottle 6 (abiotic control): test substance (30 mg) + purified water (300 mL)
Bottle 3,4,5 (abiotic control): test substance and aniline: 100 mg/L; sludge: 30 mg/L
- Measuring equipment: BOD: Closed system oxygen consumption measuring apparatus
pH: pH meter
DOC: TOC analyzer
Residual test substance amount: HPLC
- Test performed in open system: no

SAMPLING
- Sampling frequency: BOD measurement 4 times (day 7, 14, 21, 28)
- Sample storage before analysis: under nitrogen atmosphere in a desiccator at room temperature in the dark for the duration of the study

CONTROL AND BLANK SYSTEM
- Inoculum blank: Bottle 2 (inoculum blank) sludge+basal medium
- Abiotic sterile control: Bottle 6 (abiotic control): test substance (30 mg) + purified water (300 mL)
- Other:
Recovery and blank test: Duplicate samples identical to test suspension and single sample identical to inoculum blank were prepared and immediately treated by pre-analytical procedures. The amount of the residual test substance was measured to calculate the recovery rates.
Reference substance:
aniline
Key result
Parameter:
% degradation (O2 consumption)
Value:
33
Sampling time:
28 d
Details on results:
BOD in test suspension corrected with the BOD in the inoculum blank:
1: 23.3 mg
2: 22.9 mg
3: 23.4 mg

BOD in abiotic control = 0.0 mg
ThOD: maximum theoretical value was 70.6 mg O2/ 30.0mg test substance

Calculated degradabilities based on the BOD measurements:
1: 33 %
2: 32 %
3: 33 %

DOC in test suspension corrected with the DOC in the inoculum blank measured in the day 28:
1. 10.8 mg
2. 11.2 mg
3. 11.3 mg

Degradability based on the DOC:
1. 10.8 mg
2. 11.2 mg
3. 11.3 mg

DOC in abiotic control 14.9 mg (on 28 day); initial amount 17.5 mg. Degradability based on the DOC was not calculated as the DOC in abiotic control was below 90 % of the initial amount.
Key result
Parameter:
ThOD
Value:
70.6 other: mg O2/30 mg test substance

Observations of the test solution after the exposure period:

Color:

Bottle 1 (activity control): white

Bottle 6 (abiotic control): colourless

Bottle 3, 4, 5 (test suspension): colourless

Growth of the sludge: yes, in activity control and test suspensions

Validity criteria fulfilled:
yes
Interpretation of results:
inherently biodegradable, fulfilling specific criteria
Conclusions:
According to the results the test substance Incozol 4 is not readily biodegradable, but inherently biodegradable. The test substance degraded completely to water-soluble transformation products.
Executive summary:

The ready biodegradation test was conducted according to the test method relating to New Chemical Substances "Biodegradability Test of chemical Substances by Microorganisms" (Yakushokuhatsu No. 1121003, Heisei 15.11.17 Seikyoku No.3, Kanpokihatsu No.031121004, November 21, 2003, the latest revision, July 4, 2008).The test substance Incozol 4 was exposed to activated sludge in a closed-system oxygen consumption measuring apparatus.The biochemical oxygen demand (BOD) was measured over a 28-day period on every 7th day. After this period, amounts of the dissolved organic carbon (DOC) and residual test substance in test bottles were measured. The average percentage degradation based on the BOD was 33 %. This does not meet the criteria of ready degradability (≥60 %), thus, the test substance had completely disappeared, the detection of DOC at the level above 60 % suggested formation of water-soluble transformation products. The test substance was thus considered to be inherently biodegradable.

Description of key information

According to the results the test substance Incozol 4 is not readily biodegradable, but inherently biodegradable. The test substance degraded completely to water-soluble transformation products.

Key value for chemical safety assessment

Biodegradation in water:
inherently biodegradable, fulfilling specific criteria
Type of water:
freshwater

Additional information

The ready biodegradation test was conducted according to the test method relating to New Chemical Substances "Biodegradability Test of chemical Substances by Microorganisms" (Yakushokuhatsu No. 1121003, Heisei 15.11.17 Seikyoku No.3, Kanpokihatsu No.031121004, November 21, 2003, the latest revision, July 4, 2008).The test substance Incozol 4 was exposed to activated sludge in a closed-system oxygen consumption measuring apparatus. The biochemical oxygen demand (BOD) was measured over a 28-day period on every 7th day. After this period, amounts of the dissolved organic carbon (DOC) and residual test substance in test bottles were measured. The average percentage degradation based on the BOD was 33 %. This does not meet the criteria of ready degradability (≥60 %), thus, the test substance had completely disappeared, the detection of DOC at the level above 60 % suggested formation of water-soluble transformation products. The test substance was thus considered to be inherently biodegradable.