Amides, tall-oil fatty, N-[3-(dimethylamino)propyl]

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:

supporting study

Study period:

June, 29 to August, 10 1994

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:

but without effect on the validity and conclusions of the test (see details below).

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Deviations:

yes

Remarks:

but without effect on the validity and conclusions of the test (see details below).

Principles of method if other than guideline:

Devation from the OECD Guideline 301B:
As the test substance, which is relatively insoluble, is dispersed by direct emulsification in the test flasks, the CO2 is eliminated from the medium by aeration of the media after adding the substance and before introducing the inoculum.
Deaviation from the EU Method C.4-C:
The IC is measured using the MAÏHAK TOCOR 100 analyser, and its concentration is compared with the sum of the total organic carbon introduced into the mineral medium by adding the substance.That differs from the method recommended in quality plan n°5.

GLP compliance:

yes

Oxygen conditions:

aerobic

Inoculum or test system:

other: from the secondary effluent of biological urban wastewater treatment plant

Details on inoculum:

The inoculum is from the secondary effluent of the biological urban wastewater treatment plant in Versailles. The effluent, obtained on the day before seeding the flasks, is centrifuged at approximately 20ºC for 20 mins at 4000 g; the base is diluted in a volume of water in order to obtain a concentration factor of approximately 40 with respect to the sample. This inoculum is preconditioned (aeration) until the flasks are seeded. Numeration on a Totalcount Millipore total bacteria sampler shows that, for this study, the concentrated inoculum had a concentration of: 5.6* 10^6 bact/mL, which implies 5.6* 10^4 bact/mL in the final seeded medium (1% of inoculum in the incubation medium).

Duration of test (contact time):

29 d

Initial conc.:

26.7 mg/L

Based on:

test mat.

Initial conc.:

19.5 other: mg

Based on:

other: TOC/L

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

Number of culture flasks/concentration: 2 for the test medium + inoculum + test substance (Ft), 2 for blank (Fb: medium + inoculum), 1 for incolum control (Fc), 1 for sterile control (Fs) and 1 for inhibition control (Fi).
Sampling frequency: Day 0, 1, 4, 6, 8, 11, 14, 18, 21, 25, 28 and 29.
Inoculum blank: yes (test medium + inoculum).
Abiotic sterile control: yes
Inhibition control: yes (test medium + inocumum + test substance + reference substance).

After homogenisation, the parent solution of the substance is added, one drop at a time into the dilution water contained in test flasks Ft and Fi, while stirring. The reference substance is added into flasks Fc and Fi. The mercuric chloride solution is added into flasks Fs. In order to flush away the CO2, these mixtures are aired overnight with CO2 -free air. After aeration, the preconditionned inoculum is added and a sample is taken from the Ft flasks to measure the inorganic carbon, and the CO2 collectors are connected downstream of the test flaks. These collectors contain 1000 mL of barium hydroxide 0.0125M.
Incubation is in a thermostatically controlled room at 22 +/- 2°C. The air flow through the flasks is measured at the beginning of the test and whenever an upstream collector or pipe is changed.
The emanated and collected CO2 is titrated at intervals determined by the rate of progress of the reaction. The CO2 reacts with the barium dioxide to produce insoluble barium carbonate. When the white barium carbonate precipitate appears in the second collector (after the test flask), all 3 collectors are replaced by collectors containing a fresh solution of barium hydroxide. The removed collectors are titrated. On the last day of the test, after renewing the collectors, the test flasks are acidified with 1mL of concentrated hydrochloric acid in order to release the dissolved CO2. After 24hours, the quantity of the latter is measured.
The titration of the remaining barium hydroxide is carried out by neutralisation with a solution of hydrochloric acid, in the presence of phenolphthalein.

Reference substance:

benzoic acid, sodium salt

Parameter:

% degradation (CO2 evolution)

Value:

22

Sampling time:

25 d

Details on results:

The maximum degradation level is 22% in 25 days.
The latency phase for the degradation of the test substance (time between the beginning of the test and a 10% degradation) is approximately 15 days, and the percentage of the degradation after the time interval of 21 days after the latency phase is 22%.

Results with reference substance:

The percentage of degradation of the reference substance is greater than 60% in 14 days and a maximum degradation level is 65% after 25 days.

% of biodegradation in the differents series:

Days	0	1	4	6	8	11	14	18	21	25	28	29
Test	0	1	5	4	5	7	8	21	21	22	20	20
Inoculum control	0	29	58	61	62	63	63	65	65	65	63	62
Sterile	0	2	2	3	7	9	13	13	14	14	17	19
Inhibition	0	1	23	26	28	32	35	37	38	39	39	39

The substance did not reveal any inhibiting effect with respect to micro-organisms.

An abiotic sterile control was carried out: this showed that none of the substance disappears due to physical-chemical phenomena under the test conditions.

Validity criteria fulfilled:

yes

Remarks:

see details in conclusions

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

According to this test, the test substance is considered as not readily biodegradable.
Validity criteria were fulfilled:
- the inorganic carbon content in the suspension of the substance to be studied in a mineral medium (Ft flasks), at the beginning of the test (0.5mg/L), is less than 5% of the total carbon (19.5mg/L).
- the difference between the extreme values obtained for replicates with the test substance is less than 20%.
- in the toxicity test, the degradation is greater than 25% within 14d.

Executive summary:

The biodegradability study has been carried out according to the method described in the appendix C.4-C of EEC Directive 92/69/CEE. This protocol is compliant with OECD Guideline nº 301B adopted on 17/7/92.

Under the test conditions, the percentage of biodegradation of the substance was 22% in 25 days (plateau) and 20% at the end of the test (28 days).

In the time interval of 10 days after the end of the biodegradation phase, the matter reached a level of 22%.

The substance cannot be considered as readily biodegradable according to the criteria established in the OECD Guideline 301.

The criterion of validity concerning the inhibition of the test substance on micro-organisms was fulfilled (> 35%). However, some inhibition of the test substance on the reference substance was observed in the toxicity control flasks. Therefore, the low level of biodegradation of the test substance could be explained by toxicity toward microorganisms.

Under tests conditions, the test substance is assessed not readily biodegradable.

The validity criteria for the test were met.

Reference 2

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-04-07 to 2009-05-07

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:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
not applicable

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): prepared in the laboratory from secondary effluent from a wastewater treatment plant with activated sludge treating the domestic wastewater in the municipality of Abidos (France, 64).
- Concentration of sludge: between 10^7 and 10^8 cells per L
- Initial cell/biomass concentration: equivalent to a maximum level of 30 mg suspended matter/L test medium
- The inoculum has been pre-conditioned by aerating the secondary effluent, without other treatment or addition, for one day at the test temperature.

Duration of test (contact time):

28 d

Initial conc.:

2 mg/L

Based on:

test mat.

Initial conc.:

5.8 mg/L

Based on:

ThOD/L

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium:
Composition of dilution water
The dilution water was produced from the following solutions:

Solution a
8.5 g Anhydrous potassium dihydrogenophosphate (KH2PO4)
21.75 g Anhydrous potassium monohydrogenophosphate (K2HPO4)
33.40 g Dihydrated sodium monohydrogenophosphate (Na2HPO4,2H20)
Ultra-pure water made up to 1000 mL
The pH of this mixture is 7.4.

Solution b
36.40 g of dihydrated calcium chloride (CaCl2, 2 H2O) made up to 1000 mL with ultra-pure water.

Solution c
22.5 g of heptahydrated magnesium sulphate (MgSO4, 7H20) made up to 1000 mL with ultra-pure water.

Solution d
0.25 g of hexahydrated ferric chloride (FeCI3, 6H20) made up to 1000 mL with ultra-pure water. One drop of concentrated HCI is added per litre.

For 1 L of dilution water, add 800 mL of ultra-pure water, then 1 mL of each solution a to d and make up to 1000 mL with ultra-pure water.

- Additional substrate: no
- Solubilising agent (type and concentration if used): no
- Test temperature: 21 ± 2 ° C
- pH: 7.4
- pH adjusted: no, but given the buffer capacity of the mineral medium in relation to the amount of substance, no significant effect on pH is to be expected
- CEC (meq/100 g): not reported
- Aeration of dilution water: no, closed bottle test
- Continuous darkness: yes

TEST SYSTEM
- Number of culture flasks/concentration: 2
- Method used to create anaerobic conditions: use of closed, completely full BOD bottles
- Measuring equipment: oxygen specific electrode
- Test performed in closed vessels due to significant volatility of test substance: no
- Test performed in open system: no, closed bottle test
- Details of trap for CO2 and volatile organics if used: not applicable


SAMPLING
- Sampling frequency: day 0, 7, 14, 21, 28
- Sampling method: oxygen specific electrode
- Sterility check if applicable: not applicable
- Sample storage before analysis: not applicable

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes
- Abiotic sterile control: no
- Toxicity control: yes
- Positive control: yes, with sodium benzoate

STATISTICAL METHODS: no

Reference substance:

benzoic acid, sodium salt

Test performance:

The oxygen uptake of the inoculum blank has been: 0.3 mg O2/L in 28 days, which is below 1.5 mg O2/L.
The residual concentration of oxygen did not fall below 0.5 mg/L at any time.
The difference of extremes of replicate values of the removal of test chemical at the plateau, at the end of the test, or at the end of the 10-d window, was less than 20 %.
The percentage degradation of the reference item reached a level of 78% by 14 days which is above the required level of 60 %.
In the toxicity control flasks, containing both the reference item and EMULSAMINE AT-O, the percentage of biodegradation reached 71 % after 14 days. Since this value is higher than 25%, the test item is not inhibitory for inoculum.
This qualifies the test results as reliable.

Parameter:

% degradation (O2 consumption)

Value:

75

Sampling time:

28 d

Details on results:

This item reached a maximum biodegradation level of 75 % in 28 days (see Table below for details).

Results with reference substance:

The percentage degradation of the reference item (sodium benzoate) reached a level of 68 % by 14 days which is above the required level of 60 %. In the toxicity control the biodegradation of the reference substance was not significantly inhibited by the test article in a concentration of 1.8 mg/L.

Measures Of The Dissolved Oxygen By Specific Electrode

		Time (d)
Set	Vials	0	7	14	21	28
1 - Medium + inoculum
Fa	1	9.10	8.70	8.60	8.80	8.80
	2	9.10	8.70	8.70	8.80	8.80
	Mean	9.10	8.70	8.65	8.80	8.80
2 - Medium + inoculum + test substance
Ft	1	9.10	5.50	4.40	4.30	4.30
	2	9.10	5.70	4.50	4.70	4.50
	Mean	9.10	5.60	4.45	4.50	4.40
3 - Medium + inoculum + test substance + reference substance
Fi	1	9.10	5.40	4.50	4.40	4.70
	2	9.10	4.90	4.40	4.30	4.30
	Mean	9.10	5.15	4.45	4.35	4.50
4 - Medium + inoculum + reference substance
Fc	1	9.10	6.30	4.10	4.10	4.10
	2	9.10	6.90	3.80	3.80	3.90
	Mean	9.10	6.60	3.95	3.95	4.00

Table 3: Summary of results:

Specific BOD (mg of oxygen consumed by mg of substance)
Time (d):	0	7	14	21	28
Set 2 (substance)	0.00	1.55	2.10	2.15	2.20
Set 3 (control inhibition)	0.00	1.27	1.50	1.59	1.54
Set 4 (reference)	0.00	0.58	1.31	1.35	1.33
Biodegradability (average of vials)
Time (d):	0	7	14	21	28
Set 2 (substance)	0%	58%	72%	74%	75%
Set 3 (control inhibition)	0%	60%	71%	75%	73%
Set 4 (reference)	0%	35%	78%	81%	80%
Terms of Test Validity
Oxygen consumption in the set 1 < 1.5 mg/L to 28 days:					yes
Residual concentration in the sets of tests > 0.5 mg/I:					yes

Validity criteria fulfilled:

yes

Remarks:

see details in conclusions

Interpretation of results:

readily biodegradable

Conclusions:

The test item is readily biodegradable with a maximum biodegradation level of 75 % in 28 days.
The following quality criteria have been met:
The oxygen uptake of the inoculum blank has been: 0.3 mgO2/L in 28 days, which is below 1.5 mg O2/L.
The residual concentration of oxygen did not fall below 0.5 mg/L at any time.
The difference of extremes of replicate values of the removal of test chemical at the plateau, at the end of the test, or at the end of the 10-d window, was less than 20 %.
The percentage degradation of the reference item reached a level of 78% by 14 days which is above the required level of 60 %.
In the toxicity control flasks, containing both the reference item and EMULSAMINE AT-O, the percentage of biodegradation reached 71 % after 14 days. Since this value is higher than 25%, the test item is not inhibitory for inoculum.

Executive summary:

This study was performed according to the OECD guideline 301 D “Ready biodegradability - Closed Bottle Test” and GLP requirements.Under the test conditions, the percentage of biodegradation of 2 mg/L of test item reached 72 % of the ThOD after 14 days and 75 % at the end of the test (28 days). All the validity critera were fulfilled.

Reference 3

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2009-10-10 to 2009-11-15

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:

temperatrue of around 15°C during 24h in the middle of the test but without effects on the validity and conclusion of the test

Principles of method if other than guideline:

Silica was added in every flask, at a rate of 200 mg per flask.

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
not applicable

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): prepared in the laboratory from secondary effluent from a wastewater treatment plant with activated sludge treating the domestic wastewater in the municipality of Abidos (France, 64).
- Concentration of sludge: between 10^7 and 10^8 cells per L
- Initial cell/biomass concentration: equivalent to a maximum level of 30 mg suspended matter/L test medium
- The inoculum has been pre-conditioned by aerating the secondary effluent, without other treatment or addition, for one day at the test temperature.

Duration of test (contact time):

28 d

Initial conc.:

2 mg/L

Based on:

test mat.

Initial conc.:

5.8 mg/L

Based on:

ThOD/L

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium:
Composition of dilution water
The dilution water was produced from the following solutions:

Solution a
8.5 g Anhydrous potassium dihydrogenophosphate (KH2PO4)
21.75 g Anhydrous potassium monohydrogenophosphate (K2HPO4)
33.40 g Dihydrated sodium monohydrogenophosphate (Na2HPO4,2H20)
Ultra-pure water made up to 1000 mL
The pH of this mixture is 7.4.

Solution b
36.40 g of dihydrated calcium chloride (CaCl2, 2 H2O) made up to 1000 mL with ultra-pure water.

Solution c
22.5 g of heptahydrated magnesium sulphate (MgSO4, 7H20) made up to 1000 mL with ultra-pure water.

Solution d
0.25 g of hexahydrated ferric chloride (FeCI3, 6H20) made up to 1000 mL with ultra-pure water. One drop of concentrated HCI is added per litre.

For 1 L of dilution water, add 800 mL of ultra-pure water, then 1 mL of each solution a to d and make up to 1000 mL with ultra-pure water.

- Additional substrate: no
- Solubilising agent (type and concentration if used): no
- Test temperature: 21 ± 2 ° C
- pH: 7.4
- pH adjusted: no, but given the buffer capacity of the mineral medium in relation to the amount of substance, no significant effect on pH is to be expected
- CEC (meq/100 g): not reported
- Aeration of dilution water: no, closed bottle test
- Continuous darkness: yes

TEST SYSTEM
- Number of culture flasks/concentration: 2
- Method used to create anaerobic conditions: use of closed, completely full BOD bottles
- Measuring equipment: oxygen specific electrode
- Test performed in closed vessels due to significant volatility of test substance: no
- Test performed in open system: no, closed bottle test
- Details of trap for CO2 and volatile organics if used: not applicable


SAMPLING
- Sampling frequency: day 0, 7, 14, 20 and 28
- Sampling method: oxygen specific electrode
- Sterility check if applicable: not applicable
- Sample storage before analysis: not applicable

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes
- Abiotic sterile control: no
- Toxicity control: yes
- Positive control: yes, with sodium benzoate

STATISTICAL METHODS: no

Reference substance:

benzoic acid, sodium salt

Test performance:

The oxygen uptake of the inoculum blank has been: 0.65 mg O2/L in 28 days, which is below 1.5 mg O2/L.
The residual concentration of oxygen did not fall below 0.5 mg/L at any time.
The difference of extremes of replicate values of the removal of test chemical at the plateau, at the end of the test, or at the end of the 10-d window, was less than 20 %.
The percentage degradation of the reference item reached a level of 74% by 14 days which is above the required level of 60 %.
In the toxicity control flasks, containing both the reference item and the test item, the percentage of biodegradation reached 66 % after 14 days. Since this value is higher than 25%, the test item is not inhibitory for inoculum.
This qualifies the test results as reliable.

Parameter:

% degradation (O2 consumption)

Value:

60

Sampling time:

28 d

Details on results:

This item reached a maximum biodegradation level of 60% in 28 days (see Table below for details).

Results with reference substance:

The percentage degradation of the reference item (sodium benzoate) reached a level of 66 % by 14 days which is above the required level of 60 %. In the toxicity control the biodegradation of the reference substance was not significantly inhibited by the test article in a concentration of 1.8 mg/L.

Measures Of The Dissolved Oxygen By Specific Electrode

		Time (d)
Set	Vials	0	7	14	20	28
1 - Medium + inoculum
Fa	1	9.10	8.70	8.40	8.30	8.40
	2	9.10	8.70	8.50	8.40	8.50
	Mean	9.10	8.70	8.45	8.35	8.45
2 - Medium + inoculum + test substance
Ft	1	9.10	6.00	5.70	5.50	5.10
	2	9.10	6.80	5.70	5.30	4.90
	Mean	9.10	6.40	5.70	5.40	5.00
3 - Medium + inoculum + test substance + reference substance
Fi	1	9.10	5.40	4.50	4.30	4.10
	2	9.10	5.60	4.60	4.50	4.20
	Mean	9.10	5.50	4.55	4.40	4.15
4 - Medium + inoculum + reference substance
Fc	1	9.10	4.50	4.20	4.30	4.10
	2	9.10	4.40	3.80	3.80	3.70
	Mean	9.10	4.45	4.00	4.05	3.90

Table 3: Summary of results:

Specific BOD (mg of oxygen consumed by mg of substance)
Time (d):	0	7	14	21	28
Set 2 (substance)	0.00	1.15	1.38	1.48	1.73
Set 3 (control inhibition)	0.00	1.14	1.39	1.41	1.54
Set 4 (reference)	0.00	1.18	1.24	1.19	1.26
Biodegradability (average of vials)
Time (d):	0	7	14	21	28
Set 2 (substance)	0%	40%	48%	51%	60%
Set 3 (control inhibition)	0%	54%	66%	67%	73%
Set 4 (reference)	0%	71%	74%	72%	76%
Terms of Test Validity
Oxygen consumption in the set 1 < 1.5 mg/L to 28 days:					yes
Residual concentration in the sets of tests > 0.5 mg/I:					yes

Validity criteria fulfilled:

yes

Remarks:

see details in conclusions

Interpretation of results:

readily biodegradable, but failing 10-day window

Conclusions:

The test item reached a maximum biodegradation level of 60 % in 28 days. The 10-day window was not reached but its criterion is not valid to evaluate the biodegradation rates of fatty amine derivatives. Indeed, these substances are in many cases degraded by at least 2 microorganisms which results in consecutive degradation of parent and intermediate which could be visible in the degradation curve as more flattened curves. In addition, as the substance is an UVCB, this criterion can be waived.
The following quality criteria have been met:
The oxygen uptake of the inoculum blank has been: 0.3 mgO2/L in 28 days, which is below 1.5 mg O2/L.
The residual concentration of oxygen did not fall below 0.5 mg/L at any time.
The difference of extremes of replicate values of the removal of test chemical at the plateau, at the end of the test, or at the end of the 10-d window, was less than 20 %.
The percentage degradation of the reference item reached a level of 78% by 14 days which is above the required level of 60 %.
In the toxicity control flasks, containing both the reference item and EMULSAMINE AT-O, the percentage of biodegradation reached 71 % after 14 days. Since this value is higher than 25%, the test item is not inhibitory for inoculum.

Executive summary:

This study was performed according to the OECD guideline 301 D “Ready biodegradability - Closed Bottle Test” and GLP requirements. Under the test conditions, the percentage of biodegradation of 2 mg/L of test item reached 60 % of the ThOD at the end of the test (28 days). All the validity critera were fulfilled. The 10 -day window condition was not reached but since this surfactant cationic is a fatty amine derivative and an UVCB, this criterion can be waived.

Description of key information

It is assessed from the methods of manufacture the starting materials and the similarity of the end product compositions that EC272-047-7 is sufficiently similar in all respects to EC800-353-8 that it is a sensible approach to use a read-across approach. A comparison of the two substances and a read-across justification can be found in section 13 of this dataset.

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Additional information

A biodegradability study has been carried out according to the method described in the appendix C.4-C of EEC Directive 92/69/CEE (Boutonnet, 1994). This protocol is compliant with OECD Guideline nº 301B adopted on 17/7/92. Under the test conditions, the percentage of biodegradation of the substance was 20% at the end of the test (28 days). Under tests conditions, the test substance is assessed not readily biodegradable. However, two other studies have been performed according to the OECD guideline 301 D “Ready biodegradability - Closed Bottle Test” and GLP requirements (Gancet, 2009). Under the test conditions, the percentage of biodegradation of 2 mg/L of test item reached 75 % of the ThOD at the end (28 days) of the first test and 60% in the second one . All the validity criteria were fulfilled. The 10-day window condition was reached in the first test. In the second test, this criterion was not reached but since this surfactant cationic is a fatty amine derivative and an UVCB, this criterion can be waived.

The difference of biodegradation observed between these different tests can be explained by the highest test concentration (26.7 mg/L) used in the OECD 301B test compared to the OECD 301D (2 mg/L). This hypothesis is confirmed in the inhibition control of the OECD 301B test. Indeed, some inhibition of the test substance on the reference substance was observed in the toxicity control flasks.

Therefore, the low level of biodegradation of the test substance in the OECD 301B test could be explained by toxicity toward microorganisms.

To conclude, Amides, C18 -unsatd., N-[3 -(dimethylamine)propyl] was assessed to be readily biodegradable.