L-Aspartic acid, N,N'-1,2-ethanediylbis-, sodium salt (1:3)

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 26 March to 30 April 1993

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

Biodegradation was monitored in only single flasks for the control and each test substance concentration (guideline recommends duplicates). Biodegradation reached the pass level for one reference substance (aniline) but not the other (diethylene glycol).

Qualifier:

according to guideline

Guideline:

other: Directive 84/449/EEC, Brussels (1984). Part C: Methods for the Determination of Ecotoxicity: Biotic Degradation: Modified Sturm Test. Official JournaI of the European Communities L251 of 19.09.1984

Deviations:

not specified

GLP compliance:

yes

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): local (Bocholt) municipaI wastewater treatment facility receiving primarily domestic wastewater
- Preparation of inoculum for exposure: sludge was treated with a mechanical blender and allowed to settle for ±30 min. When the sludge had settled, the supernatant was decanted for use as inoculum at the rate of 10 mL/L of mineral medium.
- Initial cell/biomass concentration: number of colony forming units: 180,000/mL inoculum (measured in duplicate).

Duration of test (contact time):

ca. 34 d

Initial conc.:

10 mg/L

Based on:

test mat.

Initial conc.:

20 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: the test medium contains the following reagents per litre in high quality water (HQW, which is freshly prepared deionised water with a carbon content of 0.05 mg C/L):
4 mL of a 0.25 g/L FeCI3.6H2O solution
1 mL of a 22.5 g/L MgSO4.7H2O solution
1 mL of a 36.4 g/L CaCl2.2H2O solution
1 mL of a 40 g/L (NH4)2SO4 solution
2 mL of a phosphate solution (containing KH2PO4, K2HPO4, Na2HPO4 and NH4Cl at 8.5, 21.75, 17.69 and 1.7 g/L, respectively)
- Test temperature: 21-25°C
- pH: pH of the test suspensions at the end of the test (day 34): 7.40 (blank), 5.85 (10 mg/L test substance), 6.55 (20 mg/L test substance), 6.75 (20 mg/L aniline), 5.99 (20 mg/L diethylene glycol)


TEST SYSTEM
- Culturing apparatus: 5 L brown carboys with aeration tubes
- Number of culture flasks/concentration: one
- Method used to create aerobic conditions: test was started by bubbling CO2-free air through the suspensions at a rate of 1 to 3 bubbles per second.
- Measuring equipment: O.l.C., TOC Analyser Model 700
- Details of trap for CO2: CO2 produced in each carboy reacted with the Ba(OH)2 in the absorber bottles (125 mL absorber bottles filled with 100 mL 0.025 N Ba(OH)2) and was precipitated as BaCO3
- Other:
CO2 scrubbing apparatus: an empty glass bottle to prevent liquid carry-over, two 1 L glass bottles, one containing 700 mL of 10 N NaOH and the other 700 mL 0.025 N Ba(OH)2.8H2O, and another empty glass bottle were connected In series to a pressurised air source (synthetic air with low CO2-content). Air was sparged through the scrubbing solutions at a constant rate.

Starting-up period (purging of CO2): the preparation of the test suspensions was achieved by making dilutions directly in the CO2 test bottles:
(a) 2470 mL HQW was added to each of the test carboys.
(b) 12 mL of the FeCl3.6H2O, 6 mL of the phosphate buffer and 3 mL of each of the MgSO4.7H2O, CaCl2.2H2O and (NH4)2SO4 stock solutions were added to each carboy.
(c) 30 mL of the activated sludge inoculum was added to each carboy.
This mixture was aerated with CO2-free air for 24 hours to purge the system of CO2.

CONTROL AND BLANK SYSTEM
- Inoculum blank: 473 mL of HQW in carboy

TEST SUBSTANCE SYSTEM
- 413 mL HQW and 60 mL aniline stock solution in carboy [reference substance, final conc. 20 mg/L]
- 413 mL HQW and 60 mL diethylene glycol stock solution in carboy [reference substance, final conc. 20 mg/L]
- 413 mL HQW and 60 mL trisodium EDDS stock solution in carboy [test substance, final conc. 20 mg/L] - 433 mL HQW and 30 mL trisodium EDDS stock solution in carboy [test substance, final conc. 10 mg/L]

Performance of the test: 100 mL 0.025 N Ba(OH)2 was pipetted into each of three absorber bottles and connected in series to the exit air line of each test carboy.
The amount of CO2 produced was determined by titration of the remaining Ba(OH)2 with 0.05 N HCl. Periodically, the CO2 absorber closest to the carboy was removed for titration. The remaining two absorbers were each moved one position closer to the carboy and a new absorber containing 100 mL fresh 0.025 N Ba(OH)2 solution was placed at the far end of the series. Titrations were made before any BaCO3 precipitation was visible in the second trap.
At the end of the test the pH of the carboy content was measured and then 1 mL of concentrated HCl was added to each of the test carboys to drive off inorganic carbonate. The carboys were aerated overnight and 100 mL samples were removed for organic carbon analysis.

Reference substance:

aniline

Remarks:

20 mg/L (initial concentration)

Reference substance:

diethylene glycol

Remarks:

20 mg/L (initial concentration)

Preliminary study:

None

Test performance:

None

Parameter:

% degradation (CO2 evolution)

Value:

84.9

Sampling time:

28 d

Remarks on result:

other: 10 mg/L (initial test material concentration)

Parameter:

% degradation (CO2 evolution)

Value:

88.2

Sampling time:

35 d

Remarks on result:

other: 10 mg/L (initial test material concentration)

Parameter:

% degradation (CO2 evolution)

Value:

83.1

Sampling time:

28 d

Remarks on result:

other: 20 mg/L (initial test material concentration)

Parameter:

% degradation (CO2 evolution)

Value:

90.3

Sampling time:

35 d

Remarks on result:

other: 20 mg/L (initial test material concentration)

Details on results:

Net CO2 production and % biodegradation of the test substance: see Table 1 for details.

Results with reference substance:

- Reference substance aniline 20 mg/L (initial concentration) met the criteria for ready biodegradability (see Table 2).
- Reference substance diethylene glycol 20 mg/L (initial concentration) was substantially biodegraded but did not meet the criteria for ready biodegradability (see Table 2).

Results:

Table 1: Net CO₂ production and % biodegradation of the test substance

Day of the test	10 mg/L (initial test substance concentration)		20 mg/L (initial test substance concentration)
	mg CO2 cum. netto	biodegradation %	mg CO2 cum. netto	biodegradation %
12	7.06	19.1	6.97	9.4
14	12.65	34.2	11.31	15.3
19	23.54	63.6	35.55	48.1
23	29.83	80.6	47.65	64.4
28	31.42	84.9	61.51	83.1
35	32.63	88.2	66.81	90.3

  

Table 2: Net CO₂ production and % biodegradation of the reference substances

Day of the test	Ref. aniline 20 mg/L (initial test substance concentration)		Ref. diethylene glycol 20 mg/L (initial test substance concentration)
	mg CO2 cum. netto	biodegradation %	mg CO2 cum. netto	biodegradation %
6	49.92	30.1	1.39	1.4
10	106.77	64.4	5.54	5.6
14	126.53	76.3	12.89	13.0
28	149.71	90.2	51.41	51.9
34	151.51	91.3	62.63	63.3
35	151.93	91.6	63.23	63.9

 

Validity criteria fulfilled:

yes

Remarks:

Only single flasks were tested (guideline recommends duplicates). Only one of two reference chemicals (aniline) reached the biodegradation pass level. However, this was not critical because ready biodegradation was shown for trisodium EDDS.

Interpretation of results:

readily biodegradable

Conclusions:

A ready biodegradability test using activated sludge and carried out according to OECD Guideline 301 B (and in compliance with GLP) showed that trisodium EDDS was readily biodegradable. Initial test concentrations were 10 and 20 mg/L and biodegradation reached about 83-85% by day 28 and 88-90% by day 35.

Executive summary:

A GLP study conducted according to OECD Guideline 301 B, was performed to determine the ready biodegradability of trisodium EDDS in an aerobic aqueous medium. The substance was tested at concentrations of 10 and 20 mg/L and the inoculum was activated sludge, domestic. The test treatments, inoculum blank and the reference were measured singly (no duplicates).

At 10 mg/L, greater than 10% biodegradation of the test substance was reached on Day 12 and greater than 60% biodegradation by Day 19. The biodegradation at Day 12 was 19.1% and at Day 19 was 63.6%. Hence, the test substance met the 10-d window requirement for ready biodegradability. On Days 28 and 35, the biodegradation was 84.9% and 88.2%, respectively.

 

At 20 mg/L, greater than 10% biodegradation of the test substance was reached on Day 14 and greater than 60% biodegradation by Day 23. The biodegradation at Day 14 was 15.3% and at Day 23 was 64.4%. Hence, the test substance met the 10-d window requirement for ready biodegradability. On Days 28 and 35, the biodegradation was 83.1% and 90.3%, respectively.

 

The reference material, aniline, reached greater than 10% biodegradation on Day 6 and greater than 60% on Day 10. Hence, it met the validity criteria for a reference material.

 

In conclusion, trisodium EDDS is readily biodegradable.

Description of key information

A ready biodegradability test using activated sludge and carried out according to OECD Guideline 301 B (and in compliance with GLP) in an aerobic aqueous medium showed that trisodium EDDS was readily biodegradable.

Initial test concentrations were 10 and 20 mg/L and biodegradation reached about 83-85% by day 28 and 88-90% by day 35 (Geukens, 1993a).

In an unacclimated river water die-away test, degradation of [S,S]-EDDS reached about 75%, corresponding to a half-life of about 6.3 days, suggesting that biodegradation also occurs under realistic environmental conditions (Jaworska et al. 1999).

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Additional information

A GLP study conducted according to OECD Guideline 301 B, was performed to determine the ready biodegradability of trisodium EDDS in an aerobic aqueous medium (in a modified Sturm test). The substance was tested at concentrations of 10 and 20 mg/L and the inoculum was activated sludge, domestic. The test treatments, inoculum blank and the reference were measured singly (no duplicates). At 10 mg/L, greater than 10% biodegradation of the test substance was reached on Day 12 and greater than 60% biodegradation by Day 19. The biodegradation at Day 12 was 19.1% and at Day 19 was 63.6%. Hence, the test substance met the 10-d window requirement for ready biodegradability. On Days 28 and 35, the biodegradation was 84.9% and 88.2%, respectively. At 20 mg/L, greater than 10% biodegradation of the test substance was reached on Day 14 and greater than 60% biodegradation by Day 23. The biodegradation at Day 14 was 15.3% and at Day 23 was 64.4%. Hence, the test substance met the 10-d window requirement for ready biodegradability. On Days 28 and 35, the biodegradation was 83.1% and 90.3%, respectively. The reference material, aniline, reached greater than 10% biodegradation on Day 6 and greater than 60% on Day 10. Hence, it met the validity criteria for a reference material. In conclusion, trisodium EDDS is considered readily biodegradable (Geukens, 1993a).

In an unacclimated river water die-away test, degradation of [S,S]-EDDS was similar to that seen in the modified Sturm test, reaching 75%, corresponding to a half-life of about 6.3 days. This suggests that biodegradation also occurs under realistic environmental conditions (Jaworska et al. 1999).