N,N-bis(carboxymethyl)glycine, compound with 2-aminoethanol (1:3)

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

21/11/2005 - 27/01/2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)

Deviations:

yes

Remarks:

Degradability not measure on days 3 and 14, replaced with days 16 and 24; 4 daily temperature readings not taken.

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

natural water: freshwater

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Secondary sewage effluent from Runcorn East Sewage Works.
- Preparation of inoculum for exposure: Coarse filtered, 1 hour settling period and supernatant extraction to be used as the inoculum.
- Water filtered: yes
- Type and size of filter used, if any: coarse filtered

Duration of test (contact time):

>= 0 - <= 28 d

Initial conc.:

6.5 mg/L

Based on:

DOC

Initial conc.:

7.5 mg/L

Based on:

DOC

Parameter followed for biodegradation estimation:

DOC removal

Details on study design:

TEST CONDITIONS
- Composition of medium: nutrient enriched freshwater with sewage effluent inoculum
- Additional substrate: Positive control (reference substance, sodium benzoate
- Test temperature: 17 - 18 °C
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 272 ml glass BOD bottles
- Number of culture flasks/concentration:
- Test performed in open system: no, closed and sealed

SAMPLING
- Sampling frequency: Approximately every 7 days (0, 7, 16, 24, 28 days)

CONTROL AND BLANK SYSTEM
- Inoculum blank: Yes, Nutrient fortified freshwater only
- Toxicity control: Yes, nutrient fortified freshwater with 2.5 mg/ L sodium benzoate.

Reference substance:

benzoic acid, sodium salt

Test performance:

The reference material was degraded by 87.9% after 28 days showing that the inoculum was biologically active.

Parameter:

% degradation (DOC removal)

Value:

51

Sampling time:

7 d

Remarks on result:

other: 6.5mg/L test mat

Parameter:

% degradation (DOC removal)

Value:

70.5

Sampling time:

16 d

Remarks on result:

other: 6.5 mg/L test mat.

Key result

Parameter:

% degradation (DOC removal)

Value:

> 100

Sampling time:

28 d

Remarks on result:

other: 6.5 mg/ L test mat.

Parameter:

% degradation (DOC removal)

Value:

> 100

Sampling time:

24 d

Remarks on result:

other: 6.5 mg/L test mat.

Parameter:

% degradation (DOC removal)

Value:

48.4

Sampling time:

7 d

Remarks on result:

other: 7.5 mg/L test mat.

Parameter:

% degradation (DOC removal)

Value:

75

Sampling time:

16 d

Remarks on result:

other: 7.5 mg/L test mat.

Parameter:

% degradation (DOC removal)

Value:

> 100

Sampling time:

24 d

Remarks on result:

other: 7.5 mg/L test mat.

Parameter:

% degradation (DOC removal)

Value:

> 100

Sampling time:

28 d

Remarks on result:

other: 7.5 mg/ L test mat

Key result

Parameter:

COD

Value:

0.622 other: mg O2/mg test mat

Results with reference substance:

Sodium benzoate at 2.5 mg/ L was biodegraded to 90.3% on day 7, 86.5% on day 16, 99.2% on Day 24 and 87.9% on Day 28.

Validity criteria fulfilled:

no

Remarks:

Deviations should not have affected outcome (Biodegradability not measure on day 3 and or 14, instead measured on day 16 and 24; Temperature measurements not taken for 4 days, however not thought to have fluctuated).

Interpretation of results:

readily biodegradable

Conclusions:

The test material was biodegraded by > 100% for both 6.5 and 7.5 mg/ L on day 28.

Executive summary:

The study assesses the biodegradation of monoethanolamine salt of nitrilotriacetic acid in freshwater. The study is GLP compliant and is performed to the OECD guideline 301D: Ready Biodegradability, closed bottle method. All validity criteria were met, with the exception of the test substance interval measurements (measurements meant to be taken on days 3 and 14 were made on 14 and 24 instead) and 4 daily temperature readings were missed due to Christmas closure. However, it was concluded that these deviations did not affect the outcome of the study. The test was performed in enriched freshwater containing a secondary sewage effluent as the inoculum. The soluble reference material, sodium benzoate, degraded by more than 60% in the first 7 days, indicating that the seawater used in the test contained a satisfactory population of viable bacteria.The results from the test showed that biodegradation reached >100% at day 28 at both test concentrations of 6.5mg/L and 7.5 mg/L.

Reference 2

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

18/04/2001 - 13/06/2001

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 306 (Biodegradability in Seawater)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: ISO 6060-1989

Version / remarks:

Determination of the chemical oxygen demand

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

natural water: marine

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Collected from Biologisk stasjon, Espeland, Bergen, 40 meters depth, 10.3 °C, 50.2 mS/cm salinity, .
- Pretreatment: 4 - 14 days aging, at 20 °C, diffuse light, aeration)
- Initial cell/biomass concentration: 10 (to the power of 3) per ml bacteria

Duration of test (contact time):

>= 0 - <= 28 d

Initial conc.:

4.5 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

DOC removal

Details on study design:

TEST CONDITIONS
- Composition of medium: Nutrient fortified seawater
- Additional substrate: Inhibition control (reference substance (sodium benzoate) and test material) and reference control (reference substance (sodium benzoate)).
- Aeration of dilution water: Prior to testing
- Continuous darkness: Yes

TEST SYSTEM
- Culturing apparatus: bottles
- Number of culture flasks/concentration:
- Measuring equipment: O2 electrode
- Test performed in open system: closed

SAMPLING
- Sampling frequency: measurements taken at 7 day intervales (0, 7, 14, 21, 28 days)
- Sampling method: dissovled oxygen content measured

CONTROL AND BLANK SYSTEM
- Inoculum blank: nutrient fortified seawater
- Toxicity control: Inhibition control (reference substance (sodium benzoate) and test material) and reference control (reference substance (sodium benzoate)).

STATISTICAL METHODS: Calculation of COD according to ISO 6060-1989

Reference substance:

benzoic acid, sodium salt

Test performance:

Reference substance was biodegraded by 50% with 3-7 days showing that viable bacterial content was suitable.

Parameter:

% degradation (DOC removal)

Value:

48.2

Sampling time:

7 d

Parameter:

% degradation (DOC removal)

Value:

49.8

Sampling time:

14 d

Key result

Parameter:

% degradation (DOC removal)

Value:

54.4

Sampling time:

21 d

Parameter:

% degradation (DOC removal)

Value:

49.3

Sampling time:

28 d

Key result

Parameter:

COD

Value:

0.93 other: mg O2/ mg

Results with reference substance:

Reference substance (sodium benzoate) was tested at 2.0 mg/ L, with a ThOD of 1.665 mg O2/ mg. The reference substance biodegrade more than 60% in the first 7 days.

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

The test material was biodegraded by 10% in 2 days, 50% in 14 days and 54% in 28 days. The substance is not considered to be ready biodegradable

Executive summary:

The study assessed the aerobic biodegradability of Monoethanolamine salt of Nitrilotriacetic Acid in seawater. The study was GLP compliant and follows the OECD guideline 306: Closed Bottle Test. All validity criteria were complied with and the parameter followed to estimate ready biodegrability was dissolved oxygen content . The test was performed in nutrient fortified seawater. The results from the test showed that the registered substance biodegraded by 54.4% at day 28. This results lies close to the pass level of 60% for ready biodegradability, but it cannot be concluded that there is a potential for biodegradation in the marine environment.

Reference 3

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

13/04/2011 - 28/07/2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 306 (Biodegradability in Seawater)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

natural water: marine

Details on inoculum:

- Source of inoculum/activated sludge Natural seawater from west side of Flotta, Scapa Flow , pumped continuously from 2 metres below low water spring tide level, collection temp varies from 6 - 14 °C seasonally, 34 - 37 ‰
- Laboratory culture: Natural seawater enriched with nutrients
- Storage conditions: Stored in a 20 000 L tank, then run through 3 settlement tanks. 5-7 days before testing water moved to an aging tank, in darkness with aeration.
- Pretreatment: 45 µm filtered, nutrient enriched, sedimentation and decanting, 7 days of darkness with aeration
- Initial cell/biomass concentration: Microbial count at collection 9.40 x 10 (power of 2) CFU/ ml
- Water filtered: yes
- Type and size of filter used, if any: gravity filtered, 45 µm

Duration of test (contact time):

28 d

Initial conc.:

9.886 mg/L

Based on:

DOC

Parameter followed for biodegradation estimation:

DOC removal

Details on study design:

TEST CONDITIONS
- Composition of medium: Natural seawater enriched with nutrients: KH2PO4 8.5 g/ L, K2HPO4 21.76 g/ L, Na2HPO4.2H2O 29.92 g/ L, NH4Cl 0.5 g/ L, CaCl2 31.84 g/ L, MgSO4.7H2O 22.5 g/ L, FeCl3.6H2O 0.25 g/ L, EDTA, Di-sodium salt 0.4 g/ L
- Additional substrate: Reference substance (sodium benzoate) for positive control
- Test temperature: 19.3 - 19.9 °C
- Aeration of dilution water: prior to testing only

TEST SYSTEM
- Culturing apparatus: 270-276 ml glass BOD bottles
- Number of culture flasks/concentration: 3 replicates per time point.
- Measuring equipment: Polarographic electrode
- Test performed in open system: Closed bottle method

SAMPLING
- Sampling frequency: 7 day intervals (0, 7, 14, 21, 28 days).
- Sampling method: Measuring dissolved oxygen content
- Sample storage before analysis: 5 - 7 days prior to testing in darkness, aerated.

CONTROL AND BLANK SYSTEM
- Inoculum blank: Natural seawater enriched with nutrients (Oxygen consumption blank)
- Toxicity control: Positive control containing test material and sodium benzoate

Reference substance:

benzoic acid, sodium salt

Test performance:

The reference material (sodium benzoate) was degraded more than 60 % in the first 14 days show there was a satisfactory number of viable bacteria in the seawater

Parameter:

% degradation (DOC removal)

Value:

54

Sampling time:

7 h

Parameter:

% degradation (DOC removal)

Value:

55

Sampling time:

14 d

Parameter:

% degradation (DOC removal)

Value:

62

Sampling time:

21 d

Key result

Parameter:

% degradation (DOC removal)

Value:

88

Sampling time:

28 d

Details on results:

The test was conducted in accordance with the study plan and met all relevant validity criteria. There were no interferences in this test.

Key result

Parameter:

COD

Value:

0.526 other: mg O2/ mg

Results with reference substance:

The ready biodegradability of sodium benzoate was 69%, 84%, 78%, 65 % at 7, 14, 21, and 28 days.

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable

Conclusions:

The test material biodegraded by 88% over the 28 days (maximum biodegradation achieved) and show an inhibition of -1% to seawater bacteria

Executive summary:

The study assesses the biodegradability of N,N-bis(carboxymethyl)glycine compound with 2 -aminoethanol (1:3) in seawater. The study is GLP compliant and was performed according to the OECD guideline 306: Biodegradability in seawater. All validity criteria were met and natural seawater was used, pretreated with a 45 µm filtration, enriched with nutrients, processed via sedimentation and decanting and then aged for 7 days in darkness with 20 minutes aeration. The microbial count at collection was 9.40 x (10 (to the power of 2)) CFU/ ml and contained a satisfactory population of viable bacteria based on the reference material results. The parameter used to estimate biodegradability was dissolved oxygen content. The soluble reference material, sodium benzoate, degraded by more than 60% in the first 14 days, indicating that the seawater used in the test contained a satisfactory population of viable bacteria.The results from the test showed that biodegradation reached 88% at day 28 (3 replicates per time period).

Description of key information

The ready biodegradability of N,N'-bis(carboxymethylglycine) compound with 2 -aminoethanol (1:3) was assessed by evaluating the results from three ready biodegradability tests including a OECD 301D (key study) and two OECD 306 studies (supporting studies). The substance showed to be readily biodegradable in the OECD 301D and one of the OECD 306 tests in which biodegradation reached >100% and 88% at day 28 respectively. However, ready biodegradability was not achieved in one of the OECD 306 studies (ILAB study) in which ready biodegradability reached 54.4% at day 28. This result lies close to the pass level of 60% for ready biodegradability, but it could not be concluded that the substance showed a potential for biodegradation in the marine environment. Not all the test conditions were reported for the OECD 306 from ILAB and this is why this study received a Klimisch score of 2, but suggestion can be made on why these results led to different conclusions on the readily biodegradability of the registered substance. The OECD 306 is known to produce a large number of fails, many of which can be considered to be false negatives. A range of reasons why a fail may show during this test is that biodegradation of a chemical is dependent upon the inclusion of competent degraders within the inoculum. Low inocula concentrations such as in this test lead to the variable inclusion (and thus variable test outcome) or exclusion of specific degraders leading to a test fail. Given that N,N'-bis(carboxymethylglycine) compound with 2 -aminoethanol (1:3) has shown to be readily biodegrable in 2 of 3 OECD ready biodegradability tests and considering the variability associated with the OECD 306 test it can be concluded that the registered substance is readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Type of water:

freshwater