Reaction product of propylidynetrimethanol, propoxylated, reaction products with ammonia and 2,2-Dimethyl-3-(4-morpholinyl)propanal

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2013-05-30 to 2013-06-27

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)

Version / remarks:

May 30, 2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)

Version / remarks:

July 17, 1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 835.3110 (Ready Biodegradability)

Version / remarks:

January 1998

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Species: Activated sludge, microorganisms from a domestic waste water treatment plant.
- Origin: The (controlled) activated sludge was supplied by the sewage plant for domestic sewage in Balatonfüred, Hungary, on 30 May 2013 (on the day of the test).
- Preparation of Activated Sludge Inoculum: The activated sludge used for this study was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was re-suspended in isotonic saline solution and again centrifuged. This procedure was repeated twice. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to its dry weight was determined. Based on this ratio, calculated aliquots of washed sludge suspension, corresponding to 5 g dry material per litre was mixed with reconstituted water (see above) and then aerated until use (in this study 100 mL inoculum was prepared). Before use the sludge was filtered through cotton wool. The pH of the activated sludge inoculum after preparation was 8.11. The inoculum was not pre-adapted to the test chemical. The microbial inoculum was prepared on the day of the test.

Duration of test (contact time):

28 d

Initial conc.:

2 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Surrounding type: The test was carried out in a controlled environment room (during the formulation and oxygen measuring) at a temperature of 22 ± 2 °C according to the guideline. The test flasks were incubated in a controlled environment room at 22 ± 2 °C in the dark. Temperature was measured continuously using min/max thermometer and noticed on weekdays.
- Temperature: During the preparation, aeration and incubation of the reconstituted water and at the activated sludge inoculum preparation and incubation, the temperature was 20.4 - 22.0 °C. During the incubation (28 days) of the test units the temperature range was the following: 20.1 - 23.4 °C.
- Light Conditions: Darkness
- pH-Value of the Test Water: The pH was checked prior study start and found to be 7.58. pH adjustment was considered as not necessary.
- Oxygen Concentration of the Test Water: About 8-9 mg/L (was measured at the start of the test and found to be 8.69 mg/L)

- Preparation of the test solutions:
The respective amount, 25 mg of the test substance was weighed and suspended by mechanical dispersion (~ 10 min ultrasonic treatment under chilling) in 500 mL reconstituted water. The concentration of this test item suspension was 50 mg/L. The test item suspension was adequately stirred to ensure a good dispersion (extra care was taken for avoiding of air bubbles in the stirred solution) until use.

TEST SYSTEM
- Number of culture flasks:
10 (+2 reserve) bottles containing the test item and inoculum
10 (+2 reserve) bottles containing the reference item and inoculum (procedure control)
10 (+2 reserve) bottles containing only inoculum (inoculum control)
10 (+2 reserve) bottles containing the test item, reference item and inoculum (toxicity control)

SAMPLING
- Measurement of Oxygen: The oxygen concentration was measured with an O2 electrode [working based on LDO (Luminescent Dissolved Oxygen) method]. Oxygen measurements were performed in duplicate on days 0, 7, 14, 21 and 28.

CONTROL AND BLANK SYSTEM
- Inoculum Control: Only filtered inoculum (10 mL) was added to the aqueous test medium (5000 mL).
- Toxicity Control:Test (200 mL) and reference item (50 mL) stock solutions were mixed into the aqueous test medium (5000 mL) corresponding to the investigated test item concentration of 2 mg/L [chosen based on the preliminary information about the test item and based on its COD of the preliminary informative 3.88 mg O2/mg test item] and to 3.6 mg/L concentration of the reference item (ThODNH4 of 6.012 mg O2/L).
- Other: Microbial inoculum (2.0 mL per litre test medium) was added to each preparation bottle.

TEST PARAMETERS:
- Measurement of total oxidized N: (nitrite and nitrate):
Because of the Nitrogen content of the test item, samples for nitrate and nitrite analysis were taken from all vessels (of test item, inoculum control and toxicity control group) and the oxidized Nitrogen (nitrate and nitrite) concentrations were measured after each oxygen measurement.
- Measurement of Chemical Oxygen Demand (COD):
For determination of the COD of the test substance, test item solutions of 10 mg/L [adequately diluted from the same suspension (50 mg/L that was applied in the test] were prepared and measured. The COD of the 10 mg/L solution was determined in three parallel samples. The measured values: 17.0 mg O2/L; 17.0 mg O2/L; 16.5 mg O2/L; in average 16.8 mg O2/L that corresponds to 1.68 mg O2/mg test item.
An informative preliminary COD value was 3.88 mg O2/mg test item. At this measurement one sample was prepared in a different preparation procedure than applied in the main test.
- Measurement of temperature: Temperature was measured continuously using min/max thermometer and noticed on weekdays.

Reference substance:

benzoic acid, sodium salt

Key result

Parameter:

% degradation (O2 consumption)

Value:

-0.8

Sampling time:

28 d

Details on results:

- Correction for oxygen uptake for interference with nitrification:
Because of the N-containing test item, the oxidised nitrogen (nitrate and nitrite) concentrations were determined after each oxygen measurement with photometric method using nitrite and nitrate cell tests. The LOQ (Limit Of Quantification) of the measurements was 0.03 mg NO2/L and 0.4 mg NO3/L, respectively.
The measured quantities of nitrite and nitrate were below the LOQ (or were not detectable) in the measured samples throughout the study.
Conclusion:The BOD values were not corrected for nitrification, because the quantities of the oxidised nitrogen (nitrite and nitrate) were below of the LOQ throughout the study.

Biodegradation of Test Item:
- Percentage Biodegradation: Under the test conditions the percentage biodegradation of the test substance reached a mean of -0.8 % (that was considered as zero biodegradation) after 28 days based on its COD. (The highest percentage biodegradation value, 2.9 % was observed on the 7th day based on COD.) The slight change was within the biological variability range of the applied test system.
- Conclusion: The test item can be considered to be not ready biodegradable. According to the test guidelines the pass level for ready biodegradability is removal of 60% of COD.

Biodegradation of Toxicity Control:
- Percentage Biodegradation: In the toxicity control containing both, the test item and the reference item, a mean of 30.6 % biodegradation was noted within 14 days and a mean of 36.0 % biodegradation was determined after 28 days of incubation.
- Conclusion: According to the test guidelines the test item can be assumed as not inhibitory on the activated sludge microorganisms because the degradation in the toxicity control group was higher than 25 % within 14 days.

Results with reference substance:

The reference item Sodium benzoate was sufficiently degraded to a mean of 75.3 % after 14 days, and to a mean of 85.3 % after 28 days of incubation, based on ThODNH4. The percentage biodegradation of the reference item confirms the suitability of the used activated sludge inoculum.

Validity criteria of the Study

- Inoculum Control: The oxygen depletion in the inoculum control did not exceed 1.5 mg O2/L after 28 days. (It was 1.41 mg O2/L in average.)

- Oxygen Concentration: The residual oxygen concentration in the test flasks did not drop below 0.5 mg O2/L at any time. (The lowest value was 2.01 mg O2/L, it was measured on the 28th day in the procedure control.) The difference of duplicate values for the degradation at the plateau, at the end of the test or at the end of the 10-d window will not be greater than 20 %. Biodegradation plateau can be defined in the procedure control and in the toxicity control. The highest difference between duplicate values for degradation at these groups 2.18 %, it was observed on the 21st day. Higher differences in per cent between the duplicate values were calculated in the test item group; however the test item biodegradation was considered as zero, the obtained highest value 2.9 % on the 7th day was considered as reflect the biological variability of the test system. (The highest difference between the duplicate values at the measured oxygen concentrations was observed on the 21st day in the toxicity control, it was 3.65 %.)

- Reference item: The percentage degradation of the reference item reached the level for ready biodegradability (> 60 %) by exposure day 14. (The percentage degradation of the reference item was 75.3 % on the 14th day.)

Validity criteria fulfilled:

yes

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

Under the test conditions the percentage of ready biodegradation of the test substance reached a mean of -0.8 % (that was considered as zero biodegradation) after 28 days based on its COD. The test item can be considered to be not ready biodegradable.

Executive summary:

The ready biodegradation of the test substance was inveatigated according to EU method C.4 -E, OECD guideline 301 D and OPPTS guideline 835.3110. Under the test conditions the percentage of ready biodegradation of the test substance reached a mean of -0.8 % (that was considered as zero biodegradation) after 28 days based on its COD. (The highest percentage biodegradation value, 2.9 % was observed on the 7th day based on COD.) The slight change was within the biological variability range of the applied test system. The test item can be considered to be not ready biodegradable. The measured quantities of nitrite and nitrate were below the LOQ (or were not detectable) in the measured samples throughout the study. The reference item sodium benzoate was sufficiently degraded to a mean of 75.3 % after 14 days, and to a mean of 85.3 % after 28 days of incubation, based on ThODNH4. The percentage biodegradation of the reference item confirms the suitability of the used activated sludge inoculum.

Description of key information

Under the test conditions the percentage of ready biodegradation of the test substance reached a mean of -0.8 % (that was considered as zero biodegradation) after 28 days based on its COD. The test item can be considered to be not ready biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Type of water:

freshwater

Additional information

The ready biodegradation of the test substance was inveatigated according to EU method C.4 -E, OECD guideline 301 D and OPPTS guideline 835.3110. Under the test conditions the percentage of ready biodegradation of the test substance reached a mean of -0.8 % (that was considered as zero biodegradation) after 28 days based on its COD. (The highest percentage biodegradation value, 2.9 % was observed on the 7th day based on COD.) The slight change was within the biological variability range of the applied test system. The test item can be considered to be not ready biodegradable. The measured quantities of nitrite and nitrate were below the LOQ (or were not detectable) in the measured samples throughout the study. The reference item sodium benzoate was sufficiently degraded to a mean of 75.3 % after 14 days, and to a mean of 85.3 % after 28 days of incubation, based on ThODNH4. The percentage biodegradation of the reference item confirms the suitability of the used activated sludge inoculum.