Tetraethylammonium benzoate

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:

29-06-2016 to 27-07-2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Report date:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Tetraethylammonium benzoate

EC Number:

240-957-3

EC Name:

Tetraethylammonium benzoate

Cas Number:

16909-22-1

Molecular formula:

C8H20N.C7H5O2

IUPAC Name:

tetraethylazanium benzoate

Test material form:

liquid

Details on test material:

tetraethylammonium benzoate manufactured by Evonik Degussa GmbH, batch BRA 1434, purity 49.9 w/w (excluding solvent)

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, non-adapted

Details on inoculum:

Test system: Inoculum of the aqueous phase of non-adapted activated sludge.
Source: Municipal sewage treatment plant, 31137 Hildesheim, Germany
Reasons for the selection of the test system : Activated sludge from the sewage plant at Hildesheim is well suited as it receives predominantly municipal sewage and hardly any industrial chemical waste.
Receipt: 2016-06-23
Pretreatment: The activated sludge was washed twice with chlorine free tap water. After the second washing the settled sludge was re-suspended in mineral salts medium and was maintained in an aerobic condition by aeration for 2 hours. Thereafter the sludge was homogenized with a blender. After sedimentation the supernatant was decanted and maintained in an aerobic condition by aeration with CO2-free air for 6 days. 10 mL/L were used to initiate inoculation.
Colony forming units in the test vessels: Approx. 10E7 - 10E8 CFU/L

Duration of test (contact time):

28 d

Initial test substance concentration

Initial conc.:

52.1 mg/L

Based on:

ThOD

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

Test Item
ThOD: 1.24 mg O2/mg
ThODNO3: 1.37 mg O2/mg (with nitrification)
Test concentration: 42.0 mg/L
ThOD in the test vessel: 52.1 mg O2/L (57.5 mg O2/L with nitrification)
Pretreatment: None

Inoculum Control: Test medium without test and / or reference item

Toxicity Control: Test and reference item in test concentration

Test Method
Duration: 28 days
Application: Once at test start
Test vessels: Brown glass bottles (volume 500 mL)
Test volume: 250 mL
Test medium: Mineral salts medium according to OECD 301 F
Temperature: Nominal: 20 - 24, ± 1 °C; Actual: 20.2 – 20.5 °C
Dispersion treatment: Continuous stirring
Photoperiod: Dark, in an incubator
 
Preaparation of the test vessels: Based on the calculated oxygen demand, the test concentration of 42 mg/L, corresponding to an oxygen demand of 52.1 mg O2/L (57.5 mg O2/L with nitrification) in the vessel, was selected.

The test solutions were prepared in measuring flasks and were placed in brown glass bottles as incubation vessels (inoculum control, functional control, test item and toxicity control):
• two for the inoculum control (C1, C2)
• one for the functional control (R1)
• two for the test item (P1, P2)
• one for the toxicity control (T1)

The inoculated test medium, consisting of the required volumes of mineral medium stock solutions, ultrapure water and inoculum, was prepared in a measuring flask. 250 mL of this solution were filled in the brown glass bottles of the inoculum control, using a 250 mL measuring flask.

For the functional control, the reference item was weighed out and transferred into a measuring flask, the required vol-umes of mineral medium stock solutions, ultrapure water and inoculum, were added. 250 mL of this solution were filled in the brown glass bottle of the functional control, using a 250 mL measuring flask.
For the test item replicates, the test item was weighed out and transferred into a measuring flask, the required volumes of mineral medium stock solutions, ultrapure water and inoculum, were added. 250 mL of this solution were filled in the brown glass bottles of the test item replicates, using a 250 mL measuring flask.

For the toxicity control, the test and reference item was weighed out and transferred into a measuring flask, the required volumes of mineral medium stock solutions, ultrapure water and inoculum, were added. 250 mL of this solution were filled in the brown glass bottle of the toxicity control, using a 250 mL measuring flask.

A rubber sleeve with soda lime was hung into the opening of the bottles to absorb evolved CO2. The bottles were closed with OxiTop measuring heads and the measuring system was activated.

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

Test performance:

In the toxicity control containing both test and reference item 67% degradation occurred within 14 days. After 28 days the biodegradation came to 74%. The degradation of the reference item was not inhibited by the test item.

% Degradation

Key result

Parameter:

% degradation (O2 consumption)

Value:

36

Sampling time:

28 d

Details on results:

pH-Values at Test Start and Test End
The pH-values of the samples were measured at test start and test end (day 28).
pH-Value
Start End
Inoculum Control 7.70 1) 7.70
2) 7.67
Functional Control 7.71 7.86
Test Item 7.73 1) 7.68
2) 7.70
Toxicity Control 7.71 7.94
1), 2) = First, second replicate

BOD5 / COD results

Results with reference substance:

In order to check the activity of the test system sodium benzoate was used as functional control. The pass level > 60% was reached on day 4. The biodegradation reached a max-imum of 93 % degradation on day 24.

Any other information on results incl. tables

             Biodegradation [%] of the Test Item inComparison to the Functional Control and Toxicity Control

		Biodegradation [%]
		Study Day [d]
	Replicate	7	14	21	28
Test Item 42mg/L	1	37	41	39	39
	2	34	36	35	33
Functional Control 45 mg/L		80	89	92	92
Toxicity Control 42mg/L Test Item + 45 mg/L Reference Item		61	67	70	74

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

The test item is classified as not readily biodegradable within the 28 day period of the study.

Executive summary:

The ready biodegradability of the test item was determined with non-adapted activated sludge in the Manometric Respirometry Test for a period of 28 days. The study was conducted from 2016-06-29 to 2016-07-27 according to OECD guideline 301 F and guideline EC C.4-D atthe test facility. The test item concentration selected as appropriate was 42 mg/L, corresponding to a ThOD of 52.1 mg O2/L per test vessel. The oxygen was depleted by the respiration of bacteria and the degradation was followed by measuring the oxygen concentration. The biodegradation rate is therefore expressed as the percentage BOD (biological oxygen demand) and was calculated for each study day.

 

The mean oxygen depletion in the inoculum control was 11.2 mg O2/L on day 28.

 

In order to check the activity of the test system sodium benzoate was used as functional control. The pass level > 60% was reached on day 4. The biodegradation reached a maximum of 93 % degradation on day 24.

 

In the toxicity control containing both test and reference item 67% degradation occurred within 14 days. After 28 days the biodegradation came to 74%. The degradation of the reference item was not inhibited by the test item.

 

The biodegradation of the test item is in comparison to the readily degradable functional control and the toxicity control. Both test item replicates reached the 10% level (beginning of biodegradation) on day 2. The 60% pass level was not reached until test end and the mean biodegradation on day 28 was 36%.

The calculations were performed with the ThOD instead of the ThOD_NO3, because no indication for the typical biphasic course of biodegradation was observed and the recorded oxygen consumption resembles the oxygen demand of the benzoate moiety of the test item. Benzoate functions also as the readily biodegradable functional control and is therefore known to be easily degraded, while tertiary amines are known to be stable towards hydrolysis. Therefore, it is assumed that only the benzoate from the test item was degraded and no nitrification occurred.

The validity criteria of the guideline are fulfilled.

Conclusion

The test item is classified as not readily biodegradable within the 28 day period of the study.