N-[3-(dimethylamino)propyl]dodecanamide N-oxide

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:

19 Dec 2019 - 09 Apr 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Version / remarks:

version, 1992

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

N-[3-(dimethylamino)propyl]dodecanamide N-oxide

EC Number:

263-218-7

EC Name:

N-[3-(dimethylamino)propyl]dodecanamide N-oxide

Cas Number:

61792-31-2

Molecular formula:

C17H36N2O2

IUPAC Name:

N-[3-(dimethylamino)propyl]dodecanamide N-oxide

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: Acitvated sludge freshly otained from a municipal sewage treatment plant: 'Waterschap Aa en Maas', 's-Hertogenbosch, The Netherlands
- Storage conditions: Freshly obtained sludge was used immediately; Before the start of the test (Day -1) mineral components, Milli-RO water (ca. 80 % of final volume) and inoculum were added to each bottle. This mixture was aerated with synthetic air overnight to purge the system of CO2.
- Storage length: Freshly obtained sludge was used immediately
- Preparation of inoculum for exposure: Concentration of suspended solids (SS) was determined to be 3 g/L in concentrated sludge as used for the test. Magnetically stirred sludge was used as inoculum at an amount of 3 mL per litre of mineral medium.
- Pretreatment: Before use, sludge was coarsely sieved (1 mm)
- Concentration of sludge: 9 mg/L suspended soilds concentration

Duration of test (contact time):

29 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

118 mg/L

Based on:

test mat.

Initial test substance concentration 2

Initial conc.:

12 mg/L

Based on:

TOC

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: According to the guideline
- Test temperature: 22 - 23 °C
- pH: 7.4 - 7.5 (Control and Test item concentrations)
- Aeration of dilution water: Approximately 1-2 bubbles per second (ca. 30-100 mL/min).
- Continuous darkness: Yes, Test media were excluded from light.

TEST SYSTEM
- Culturing apparatus: 2 litre brown coloured glass bottles.
- Number of culture flasks/concentration: 2 bottles
- Method used to create aerobic conditions: A mixture of oxygen (ca. 20%) and nitrogen (ca. 80%) was passed through a bottle; Synthetic air was passed through the scrubbing solutions.
- Measuring equipment: CO2 produced in each test bottle reacted with barium hydroxide in the gas scrubbing bottle and precipitated out as barium carbonate. The amount of CO2 produced was determined by titrating remaining Ba(OH)2 with 0.05 M standardized HCl (1:20 dilution from 1 M HCl (Titrisol® ampoule), Merck, Darmstadt, Germany).
- Test performed in open system: Yes
- Details of trap for CO2 and volatile organics if used: Three CO2-absorbers (bottles filled with 100 mL 0.0125 M Ba(OH)2) were connected in series to the exit aeration line of each test bottle.

SAMPLING
- Sampling frequency: Every second or third day during the first 10 days, and thereafter at least every fifth day until Day 28, for inoculum blank and test item. Titrations for procedural and toxicity control were made over a period of
at least 14 days. Final titration was made on Day 15 (procedural and toxicity control) and on Day 29 (remaining vessels).
- Sampling method: Each time the CO2-absorber nearest to the test bottle was removed for titration; each of the remaining two absorbers were moved one position in the direction of the test bottle. A new CO2-absorber was placed at the far end of the series. On the penultimate day, pH of respective test suspensions was measured and 1 mL of concentrated HCl (37%, Merck) was added to the inoculum blank and test suspension.

CONTROL AND BLANK SYSTEM
- Inoculum blank: 2 bottles
- Toxicity control: 1 bottle
- Reference control: 1 bottle

Reference substance

Reference substance:

acetic acid, sodium salt

Results and discussion

% Degradation

% Degradation 1

Parameter:

% degradation (CO2 evolution)

Value:

74

Sampling time:

28 d

Remarks on result:

other: based on ThCO2

Remarks:

Bottle A

% Degradation 2

Parameter:

% degradation (CO2 evolution)

Value:

65

Sampling time:

28 d

Remarks on result:

other: based on ThCO2

Remarks:

Bottle B

BOD5 / COD results

Results with reference substance:

In the toxicity control, more than 25% biodegradation occurred within 14 days (72%, based on ThCO2). Therefore, the test item was considered not to inhibit microbial activity.

Any other information on results incl. tables

Procedure control (reference control):

Functioning of the test system was checked by testing the procedural control item sodium acetate, which showed a normal biodegradation curve.

Conclusion:

Relative biodegradation values calculated from measurements performed during the test period revealed 74% and 65% biodegradation of the test item (based on ThCO2). Thus, the criterion for ready biodegradability (at least 60% biodegradation within a 10-day window) was met.

Table 1: CO2 Production and Biodegradation of Test Item (Bottle A)

Day	HCl (0.05 N) titrated (mL)		Produced CO2	Produced CO2	Cumulative CO2	Biodegradation 1)
	Blank (mean)	Bottle A	(mL HCl)	(mg)	(mg)	(%)
2	44.73	41.44	3.29	3.6	3.6	4
5	43.00	23.70	19.30	21.2	24.8	29
8	41.57	26.38	15.19	16.7	41.6	48
12	40.53	30.62	9.91	10.9	52.4	61
15	42.23	38.46	3.77	4.1	56.6	66
19	41.59	39.43	2.16	2.4	59.0	69
23	41.34	39.88	1.45	1.6	60.6	70
29 2)	40.74	39.31	1.43	1.6	62.1	72
29 2)	40.87	39.24	1.63	1.8	63.9	74
29 2)	42.76	43.45	0.00	0.0	63.9	74

1): Calculated as the ratio between CO2 produced (cumulative) and the ThCO2 of the test item: 86.0 mg CO2/2L.

2): CO2 measured on Day 29 is actually part of CO2 production of Day 28, since microbial activity was ended on Day 28 by addition of HCl.

 

Table 2: CO2 Production and Biodegradation of Test Item (Bottle B)

Day	HCl (0.05 N) titrated (mL)		Produced CO2	Produced CO2	Cumulative CO2	Biodegradation 1)
	Blank (mean)	Bottle B	(mL HCl)	(mg)	(mg)	(%)
2	44.73	42.03	2.70	3.0	3.0	3
5	43.00	25.30	17.70	19.5	22.4	26
8	41.57	26.62	14.95	16.4	38.9	45
12	40.53	29.74	10.79	11.9	50.7	59
15	42.23	38.75	3.48	3.8	54.6	63
19	41.59	40.98	0.61	0.7	55.2	64
23	41.34	41.31	0.02	0.0	55.3	64
29 2)	40.74	39.49	1.25	1.4	56.6	65
29 2)	40.87	42.10	0.00	0.0	56.6	65
29 2)	42.76	43.99	0.00	0.0	56.6	65

1): Calculated as the ratio between CO2 produced (cumulative) and the ThCO2 of the test item: 86.5 mg CO2/2L.

2): CO2 measured on Day 29 is actually part of CO2 production of Day 28, since microbial activity was ended on Day 28 by

addition of HCl.

 

Table 3: CO2 Production and Biodegradation in Toxicity Control

Day	HCl (0.05 N) titrated (mL)		Produced CO2	Produced CO2	Cumulative CO2	Biodegradation1)
	Blank (mean)	Toxicity control	(mL HCl)	(mg)	(mg)	(%)
2	44.73	31.72	13.01	14.3	14.3	8
5	43.00	6.30	36.70	40.4	54.7	32
8	41.57	11.36	30.21	33.2	87.9	51
12	40.53	19.65	20.88	23.0	110.9	64
152)	42.23	29.57	12.66	13.9	124.8	72

1): Calculated as the ratio between CO2 produced (cumulative) and the sum of the ThCO2 of the test item and procedural control item: 172.6 mg CO2/2L (ThCO2 test item: 86.4 mg CO2/2L +ThCO2 sodium acetate: 86.2 mg CO2/2L).

2): CO2 measured on Day 15 is actually part of CO2 production of Day 14, since microbial activity was ended on Day 14 by addition of HCl.

 

Table 4: Comparison of Biodegradation of Test Item in Bottles A and B

Day	Biodegradation (%)
	Bottle A	Bottle B	Mean A and B	Δ A-B 1)
2	4	3	4	1
5	29	26	27	3
8	48	45	47	3
12	61	59	60	2
15	66	63	64	3
19	69	64	66	5
23	70	64	67	7
29 2)	72	65	69	7
29 2)	74	65	70	9
29 2)	74	65	70	9

1): Absolute difference in biodegradation between bottles A and B

2): Biodegradation is ended on Day 28 by addition of HCl. Therefore, differences observed on Day 29 are actually differences of Day 28.

Table 5: Validity criteria

Criterion from the guideline	Outcome	Validity criterion fulfilled
Difference of extremes of replicate values of the removal of the test chemical at the plateau, at the end of the test or at the end of the 10-d window, as appropriate, is less than 20%.	≤9%	Yes
Percentage degradation of the reference compound reached the pass level by day 14 (≥ 60%).	78%	Yes
The toxicity control should degrade to at least 35% (based on DOC) or at least 25% (based on ThOD or ThCO2) within 14 d.	72% based on ThCO2	Yes
The IC content of the test substance suspension in the mineral medium at the beginning of the test must be less than 5% of the TC.	Inorganic Carbon content (IC) of test item in mineral medium at the beginning of the test was less than 5 % of the Total Carbon content (TC).	Yes
The total CO2 evolution in the inoculum blank at the end of the test should not normally exceed 40 mg/L medium.	33.1 mg CO2/L	Yes

 

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”

Interpretation of results:

readily biodegradable