N-[2-[(4-amino-m-tolyl)ethylamino]ethyl]methanesulphonamide

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:

31 Oct - 30 Nov 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

N-(2-(4-amino-N-ethyl-m-toluidino)ethyl)methanesulphonamide sesquisulphate

EC Number:

247-161-5

EC Name:

N-(2-(4-amino-N-ethyl-m-toluidino)ethyl)methanesulphonamide sesquisulphate

Cas Number:

25646-71-3

Molecular formula:

C12H21N3O2S.3/2H2O4S

IUPAC Name:

Methanesulfonamide, N-[2-[(4-amino-3-methylphenyl)ethylamino]ethyl]-, sulfate (2:3)

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

other: supernatant of activated sludge, non-adapted, domestic

Details on inoculum:

- Source of inoculum/activated sludge: Activated sludge freshly obtained from a municipal sewage treatment plant: 'Waterschap Aa en Maas', 's-Hertogenbosch, The Netherlands, receiving predominantly domestic sewage. The supernatant liquid was used as inoculum.
- Storage conditions: Mineral components, Milli- RO water and inoculum was aerated with synthetic air overnight to purge the system of CO2.
- Storage length: Over night
- Pretreatment: The freshly obtained sludge was kept under continuous aeration until further treatment. Before use, the sludge was allowed to settle (41 min) and the supernatant liquid was then used as inoculum at the amount of 10 mL/L of mineral medium.
- Concentration of sludge: 4.3 g/L in the concentrated sludge.

Duration of test (contact time):

28 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

36.5 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 guideline; tap-water purified by reverse osmosis (Milli- RO) and subsequently passed over activated carbon
- Test temperature: 22 - 23 °C
- pH: 7.5 - 7.9
- pH adjusted: yes
- Aeration of dilution water: aerated and stirred continuously at a rate of approximately 1-2 bubbles per second (ca. 30-100 mL/min).
- Suspended solids concentration: 4.3 g/L in the concentrated sludge
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 2 litre brown coloured glass bottles
- Number of culture flasks/concentration: 2 bottles (with test item)
- Method used to create aerobic conditions: A mixture of oxygen (ca. 20%) and nitrogen (ca. 80%) was passed through a bottle, containing 0.5 - 1 litre 0.0125 M Ba(OH)2 solution to trap CO2 which might be present in small amounts. The synthetic air was passed through the scrubbing solutions at a rate of approximately 1-2 bubbles per second (ca. 30-100 mL/min).
- Measuring equipment: The CO2 produced in each test bottle reacted with the barium hydroxide in the gas scrubbing bottle and precipitated out as barium carbonate. The amount of CO2 produced was determined by titrating the 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 closed vessels: no
- Details of trap for CO2: emitted CO2 was trapped in 0.25 M NaOH. Two scrubbers containing 100 mL each were connected in series to the test vessels. The initial IC value of the 0.25 M NaOH was separately determined in each flask

SAMPLING
- Sampling frequency: Titrations were made every second or third day during the first 10 days, and thereafter at least every fifth day until day 28, for the inoculum blank and test item. Positive and toxicity control: at laest 14 days
- Sampling method: The CO2 produced in each test bottle reacted with the barium hydroxide in the gas scrubbing bottle and precipitated out as barium carbonate. The amount of CO2 produced was determined by titrating the remaining Ba(OH)2 with 0.05 M standardized HCl (1:20 dilution from 1 M HCl (Titrisol® ampoule), Merck, Darmstadt, Germany).

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

Reference substance

Reference substance:

acetic acid, sodium salt

Results and discussion

% Degradation

% Degradation 1

Parameter:

% degradation (CO2 evolution)

Value:

11

Sampling time:

28 d

Remarks on result:

other: Vessel A

% Degradation 2

Parameter:

% degradation (CO2 evolution)

Value:

20

Sampling time:

28 d

Remarks on result:

other: Vessel B

BOD5 / COD results

Results with reference substance:

The positive control item was biodegraded by at least 60% (73%) within 14 days.

Any other information on results incl. tables

Biological Results:

Table 1: CO2 Production and Percentage Biodegradation of the Positive Control Item

Day	HCl (0.05 N) titrated (mL)		Produced CO2	Produced CO2	Cumulative CO2	Biodegradation 1)
	Blank (mean)	Positive Control	(mL HCl)	(mg)	(mg)	(%)
2	47.08	45.21	1.87	2.1	2.1	2
5	46.19	22.01	24.18	26.6	28.6	33
8	45.69	32.09	13.60	15.0	43.6	51
12	45.65	35.41	10.24	11.3	54.9	64
152)	46.23	39.14	7.09	7.8	62.7	73

1): Calculated as the ratio between CO2 produced (cumulative) and the ThCO2 of sodium acetate: 85.8 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 2: CO2 Production and Percentage Biodegradation of the Toxicity Control

Day	HCl (0.05 N) titrated (mL)		Produced CO2	Produced CO2	Cumulative CO2	Biodegradation 1)
	Blank (mean)	Toxicity control	(mL HCl)	(mg)	(mg)	(%)
2	47.08	47.10	0.00	0.0	0.0	0
5	46.19	24.46	21.73	23.9	23.9	14
8	45.69	26.14	19.55	21.5	45.4	26
12	45.65	33.67	11.98	13.2	58.6	34
152)	46.23	35.95	10.28	11.3	69.9	40

1): Calculated as the ratio between CO2 produced (cumulative) and the sum of the ThCO2 of the test item and positive control: 173.8 mg CO2/2L (ThCO2 test item: 88.0 mg CO2/2L + ThCO2 sodium acetate: 85.8 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 3: Comparison of Biodegradation of the Test Item in Bottles A and B

Day	Biodegradation (%)
	Bottle A	Bottle B	Mean A and B	Δ A-B 1)
2	0	1	1	1
5	1	2	2	1
8	3	4	4	1
12	6	6	6	0
15	7	10	9	3
19	8	13	11	5
23	9	16	13	7
29 2)	11	19	15	8
29 2)	11	20	16	9
29 2)	11	20	16	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 3: Validity criteria for OECD 301 B.

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 has reached the pass levels by day 14.	73%	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.	40%	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.	< 5%	Yes
The total CO2 evolution in the inoculum blank at the end of the test should not normally exceed 40 mg/L medium.	38.6 CO2 /2 L 19.3 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:

not readily biodegradable

Conclusions:

In conclusion, the test item was not readily biodegradable under the conditions of the modified Sturm test presently performed.