N,N'-hexamethylenebis(cinnamylideneamine)

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:

From July 06 to August 04, 2016

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Version / remarks:

1997

Qualifier:

according to guideline

Guideline:

EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)

Version / remarks:

2008

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, non-adapted

Details on inoculum:

Non-adapted activated sludge from the aeration tank of the ARA Werdhölzli (CH-8048 Zürich), a municipal biological waste water treatment plant. 30 mg/l dry matter in the final mixture.

Duration of test (contact time):

ca. 28 d

Initial conc.:

ca. 32.8 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

Test procedure
The activated sludge was used after sampling from the treatment plant without adaptation. However, the sludge was pre-conditioned for 7 days (aerated but not fed) to reduce the amount of O2 consumed by the blank controls. For this pre-conditioning phase, the sludge was washed twice with tap water and once with mineral medium. After centrifugation, the sludge was suspended in test medium, at about 1.8 g/l dry matter. Before the test, this suspension was diluted down to 60 mg/l dry matter, i.e. twice the final concentration, since this suspension was diluted 1:1 (v:v) afterwards (either with mineral medium in flasks T and B; or with the reference item’s stock solution in flasks R and X, or with ultrapure water in flask C).
The test item was applied by direct addition in the test vessels, so as to give a final test concentration of about 100 mg ThOD/l (i.e. about 32 mg/l test item).
The reference item was dissolved in the test medium at a concentration twice of the final concentration to be achieved for the test. This stock solution was diluted with the sludge suspension 1:1 (v:v) to give a final test concentration of about 100 mg ThOD/l (i.e. about 60 mg/l reference item).
In the toxicity control a mixture of about 100 mgThOD/l of test item and about 100 mg ThOD/l of reference item was tested (total 200 mg ThOD/l; application by direct addition and by stock solution, respectively).
In the abiotic sterile control about 100 mg ThOD/l of test item was tested (application by direct addition).
The following test flasks was set up:
T: Test suspension: containing inoculum, test medium and test item (two replicates)
B: Blank control: containing inoculum and test medium (two replicates)
R: Procedure control containing inoculum, test medium and sodium benzoate as ready biodegradable reference item (two replicates)
C: Abiotic sterile control: containing test item, ultra-pure water and 0.04 mM HgCl2 as sterilizing agent to prevent microbial decomposition (one replicate)
X: Toxicity control: containing inoculum, test medium, test item and sodium benzoate as ready biodegradable reference item (one replicate)
Before the test start, the pH value was checked and if necessary adjusted to 7.2 (± 0.2) with NaOH or HCl (except in flask C; and in flask T and X only if the test item is soluble).
The test vessels were stirred by an inductive stirring system for a maximum test period of 28 days. During the test the O2 uptake was continuously measured with a manometric BOD measuring device. The temperature was recorded with a data logger.
At the end of the test, the pH was measured in all flasks except the abiotic sterile control (C).
Total elimination was determined in each test vessel (T, B and R) based on the theoretical carbon content and the analysis of the dissolved organic carbon (DOC) concentrations at the end of the test.

Reference substance:

benzoic acid, sodium salt

Parameter:

% degradation (O2 consumption)

Value:

ca. 77

Sampling time:

10 d

Remarks on result:

other: 10-d window

Key result

Parameter:

% degradation (O2 consumption)

Value:

ca. 90

Sampling time:

28 d

Remarks on result:

other: as compared to the theoretical oxygen demand (ThOD).

Details on results:

Biodegradation of the test item started without any significant lag-phase.
The procedure control sodium benzoate reached 80 % biodegradation after 14 days, thus confirming suitability of inoculum and test conditions.
The respective concentrations of organic carbon at the beginning (as mg TOC/l) and at the end (as mg DOC/l) of the test after 28 days were, respectively:
27.4 * and 1.24 / 1.66 for test unit 1 / test unit 2
<0.5 and 0.776 for the blank control (mean of two replicates)
35.0* and 1.32 for the procedure control (mean of two replicates)
* theoretical value calculated based on the carbon content and the test concentration

The total eliminationcalculated based on dissolved organic carbon (DOC) measurements reached 98 % for the test item and for sodium benzoate, respectively. The data confirm the high biodegradability of the test item since the total elimination based on DOC was only slightly higher than the biodegradation based on BOD.
The test item reached the pass level of 60% for ready biodegradability in the Manometric Respirometry Test within the 10-d window

Additional controls

Toxicity control

At the applied initial test concentration of 32.5 mg/l, the test item was not judged to have any inhibitory effect on the microbial population, since the biodegradation of the mixture (test item + reference item sodium benzoate) exceeded 25 % within 14 days.

Abiotic sterile control

The test item was not abiotically degraded (by processes using O₂) during the whole test period of 28 days in the absence of microorganisms as confirmed by the lack of oxygen consumption.

Validity of the test

All validity criteria were fulfilled.

At the end of the test the pH values of the blank controls were 7.7 and 7.2 and the pH values of the procedure control were both 7.4. The pH values of the test units were 7.2 and 7.1, and the pH of the toxicity control was 7.3. All pH values were therefore in the required range at the end of the test.

The temperature was in the acceptable range during the whole incubation period.

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable

Conclusions:

The test substance under these test conditions is Readily Biodegradable.

Executive summary:

Method

The biodegradability of the test substance exposed to microorganisms derived from activated sludge of a municipal sewage treatment plant was investigated under aerobic static exposure conditions, following the test guideline OECD 301 F.

In this test, biodegradability of the test item is monitored during 28 days at 22 °C in test vessels containing a low concentration of sludge from a sewage treatment plant (30 mg/l dry matter), test medium and the test item (about 100 mg ThOD /l); along with blank (negative), procedure (positive), toxicity and abiotic sterile controls. The amount of O₂ taken up by the microbial population during biodegradation of the test item (corrected for the value in the blank control) is expressed as percentage of the theoretical O₂demand, ThOD, and serves as the measure for the extent of biodegradation.

Additionally, total elimination was determined based on the theoretical carbon content and the analysis of the dissolved organic carbon (DOC) concentrations at the end of the test.

Results

The biodegradability of the test substance based on O₂consumption was calculated to be 90 % after 28 days as compared to the theoretical oxygen demand (ThOD).

The biodegradation reached 77 % at the end of the 10-d window.

Biodegradation of the test item started without any significant lag-phase.

The procedure control sodium benzoate reached 80 % biodegradation after 14 days, thus confirming suitability of inoculum and test conditions.

The total eliminationcalculated based on dissolved organic carbon (DOC) measurements reached 98 % for the test item and for sodium benzoate, respectively. The data confirm the high biodegradability of the test item since the total elimination based on DOC was only slightly higher than the biodegradation based on BOD.

The test substance reached the pass level of 60 % for ready biodegradability in the Manometric Respirometry Test within the 10-d window.

Conclusion

The test substance is Readily Biodegradable.

Description of key information

Readily Biodegradable

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Additional information

The biodegradability of the test substance was evaluated according to the test guideline OECD 301 F.

In this test, biodegradability of the test item is monitored during 28 days based on ThOD. Additionally, total elimination was determined based on the theoretical carbon content and the analysis of the dissolved organic carbon (DOC) concentrations at the end of the test.

The test substance reached the pass level of 60 % for ready biodegradability in the Manometric Respirometry Test within the 10-d window and reached the 90 % of biodegradation after 28 days, therefore is considered as readily biodegradable.