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Environmental fate & pathways

Biodegradation in water: screening tests

Administrative data

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
29 December 2018 - 22 February 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report date:
2019

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
Octadecanoic acid, reaction products with diethylenetriamine, di-Me sulfate-quaternized
EC Number:
291-707-5
EC Name:
Octadecanoic acid, reaction products with diethylenetriamine, di-Me sulfate-quaternized
Cas Number:
90459-62-4
Molecular formula:
C24H55N3O6S
IUPAC Name:
bis(2-aminoethyl)amine octadecanoic acid dimethyl sulfate
Test material form:
solid
Remarks:
paste

Study design

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): A fresh sample of activated sludge was collected from the aeration tank of a sewage treatment plant unit treating predominantly domestic sewage at Siddaganga Institute of Technology, Chandana Complex, Tumkur – 572103, India.
The coarse particles were removed by filtration and the concentrated sludge was suspended in mineral medium to yield a concentration of approximately 3-5 g suspended solids/L. After complete re-suspension was achieved, a sample was withdrawn for the determination of the dry weight of the suspended solids. A well-mixed volume of 100 mL of an activated sludge was filtered through a pre-weighed crucible. The crucible was heated to constant mass at 105ºC for 1 hour and then weighed. The suspended solids were determined to be 3325 mg/L and aerated for pre-conditioning the inoculum until inoculation at the test temperature.
Accurately 27.1 mL of the inoculum were added to each of the flasks to attain a concentration of 30 mg/L of suspended solids.
Duration of test (contact time):
28 d
Initial test substance concentrationopen allclose all
Initial conc.:
14.2 mg/L
Based on:
test mat.
Initial conc.:
15 mg/L
Based on:
TOC
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST CONDITIONS
The test solutions were prepared and inoculated in 5 L test culture vessels each containing 3 L of solution.
Accurately 24 hours prior to addition of the test and reference items, the vessels were filled with 2400 mL of mineral medium and 27.1 mL of the activated sludge was added to get a final concentration of suspended solids of 30 mg/L in 3 L of inoculated mixture. The vessels were aerated overnight at test temperature. On Day 0, the test and reference substance stock solutions were added and the pH of all the vessels was measured using a calibrated pH meter. The pH values of all the vessels were within 7.4 ± 0.2 before being adjusted to 3 L by the addition of mineral medium and continuously purged using CO2 free air. The vessels contents were continuously flushed for 29 days by bubbling CO2 free air at a rate of 30 - 100 mL/minute. The flow of the air was checked once per day on all working days.

SAMPLING
- Sampling frequency: During the first ten days, the analysis of CO2 was performed every third day and then at least every fifth day until the 28th day.
The air outlet from each vessel was connected to three absorption bottles in series, filled with 100 mL of 0.0125 M Ba(OH)2 solution and connected in series to the exit air line of each 5 L vessel. The CO2 produced in each experimental vessel reacts with the Barium hydroxide in CO2 adsorption bottles and precipitated as barium carbonate. The amount of CO2 produced was determined by titrating the remaining Ba(OH)2 with 0.05 N HCl. On the day of CO2 measurement, the barium hydroxide absorber closest to the test vessel for titration was disconnected. The remaining absorbers were moved one place closer to the test vessel and a new absorber containing
100 mL fresh 0.0125 M barium hydroxide was placed at the far end of the series. The volume of Ba(OH)2 solution in the CO2 adsorption bottle was measured and transferred to a 250 mL volumetric flask. The Ba(OH)2 was then titrated with 0.05 N HCl using phenolphthalein as an indicator and a stirring bar and magnetic stirrer for good mixing. The titration was completed when the indicator became colorless.
On day 28 of the study, the pH of each test vessel was measured and recorded. A volume of 1 mL of concentrated HCl was added to each of the vessels, which was then aerated overnight to remove the CO2 present in the test suspensions. On the next day, day 29, the last analysis of evolved carbon dioxide was performed. The samples were taken from the CO2 absorber vessels at pre-determined intervals of 3, 6, 9, 14, 19, 24, 28 and 29 days and analyzed for CO2 immediately.
The amount of CO2 produced by the test substance during the test was measured and expressed as percent of the theoretical CO2 (% of ThCO2) calculated from the TOC content of the test compound added. Biodegradability was expressed as percentage ThCO2.

CONTROL AND BLANK SYSTEM
- Inoculum blank: 2 replicates
- Procedure Control: 2 replicates


Reference substance
Reference substance:
acetic acid, sodium salt

Results and discussion

% Degradation
Parameter:
% degradation (CO2 evolution)
Value:
69.1
Sampling time:
28 d
Remarks on result:
other: UVCB substance , thus 10 d window criterion does not apply
Details on results:
The test item reached a maximum mean biodegradation value of 69.10% (% of ThCO2) for two bottles within 28 days exceeding the criterion of 60% biodegradation given by the guideline.
The total CO2 evolution in the Control Blanks at the end of the test was 38.12 mg/L and does not exceed 40 mg/L.
The differences in replicate values of the removal of the test substance at the end of the test were less than 20% (i.e -0.77%).

BOD5 / COD results

Results with reference substance:
The degradation of the procedure control substance (Sodium acetate) attained 63.74% (% of ThCO2) biodegradation by 14 days (greater than 60% degradation).

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The results of the CO2 Evolution Test indicated that Octadecanoic acid, reaction products with diethylenetriamine, di-Me sulfate-quaternized is readily biodegradable according to OECD criteria. The test item is an UVCB substance and is readily biodegradable after 28 days (69.10% of ThCO2) under the conditions of the CO2 Evolution test.
Executive summary:

To assess the ready biodegradability of the test item Octadecanoic acid, reaction products with diethylenetriamine, di-Me sulfate-quaternized a GLP-compliance CO2 Evolution Test (Modified Sturm Test) according to OECD TG No. 301 B (adopted 17th July 1992) was carried out.

During the study, the amount (mg) of CO2 produced was calculated at the start of the experiment and on days 3, 6, 9, 14, 19, 24 and 28, respectively. The relative biodegradation values were calculated from the CO2 measurements during the test period of 28 days.

The test item reached a maximum mean biodegradation value of69.10%(% of ThCO2) for two bottles within 28 days exceeding the criteria of 60% biodegradation given by the guideline.

The total CO2 evolution in the Control Blanks at the end of the test was 38.12 mg/L which is below the maximum of 40 mg/L defined in the validity criteria.

The differences in replicate values of the removal of the test item at the end of the test were less than 20% (i.e -0.77%). 

The percentage degradation of the procedure control substance (Sodium acetate) attained 63.74% (% of ThCO2) biodegradation after 14 days(greater than 60% degradation).

The CO2 absorber traps were titrated on day 29 after acidifying the test flasks on day 28. This acidification drives off any dissolved CO2 present in the test vessels. The results of the 0.05 N HCl consumed for all the test flasks from the absorber vessels on Day 29 showed no significant changes in any of the test flasks as compared to the blank BaOH2 and confirmed that no significant production of CO2 into the absorber vessels occurred after acidifying the test flasks.

All the criteria for acceptability of the test were met. Hence, the present study was considered to be valid.

The results of the CO2 Evolution Test indicated that the test item is readily biodegradable according to OECD criteria. The test item is an UVCB substance and is readily biodegradable after 28 days (69.10% of ThCO2) under the conditions of the CO2 Evolution test.