Propanol, iminobis-, N-(C16-18 and C18-unsatd. alkyl) derivs.

Biodegradation in water and sediment: simulation tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: sewage treatment simulation testing

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

08-09-2010 - 29-11-2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

See section 13.2 for the read-across justification.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 303 A (Simulation Test - Aerobic Sewage Treatment. A: Activated Sludge Units)

Deviations:

yes

Remarks:

acceptable deviations

Principles of method if other than guideline:

A few minor deviations to the guidelines were introduced. The primary settled sewage was collected weekly and stored in the refrigerator until required instead of a daily collection of wastewater. The units consisted of aeration vessels capable of holding only 0.35 L from which the liquor was then passed continuously to settler of 0.30 liter capacities.

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

no

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

Secondary activated sludge to inoculate the test at the start was collected on 08-09-2010 from the wastewater treatment plant (WWTP) Nieuwgraaf in Duiven, The Netherlands. The WWTP Nieuwgraaf is an activated sludge plant treating predominantly domestic sewage. 0.35 liter of secondary activated sludge containing approximately 3 g/L dry weight was used as an inoculum for each CAS unit. This dry weight was obtained by diluting the sludge
obtained from the treatment plant. To maintain an activated sludge concentration between 2.0 and 3.0 g/L secondary activated sludge containing
approximately 3 g/L dry weight was collected weekly. The primary settled sewage was collected from the sameplant weekly and stored frozen until
required. Before use 10 mL/L of a NaHCO3 (10 g/L) solution was added.

Duration of test (contact time):

62 d

Initial conc.:

28.2 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

DOC removal

Details on study design:

The CAS test was performed in Husmann-type units constructed of glass. The units consisted of an aeration vessel capable of holding 0.35 liter fromwhich the liquor was passed continuously to a settler of 0.3 liter. The domestic waste water liquor in a cooled vessel was supplied with a pump. The liquor passed through the aeration vessel and settler and treated effluent left the apparatus to be collected in a vessel. Aeration was achieved through a capillary on the bottom of the aeration section at a rate of approximately 8 L/h of air. Sludge accumulating around the top of the aeration vessel was returned in the system once a day by brushing.

A suspension of ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N- (hydrogenated tallow alkyl) derivatives of 4.1 g/L in deionized water was directly added to the test unit using a syringe pump. The stock was prepared by adding 4.1 g of test substance to 0.9 L of deionized water. A
homogenous suspension was obtained by acidifying the stock to a pH of approximately 7 by adding HCl.
The final stock suspension was made up to 1.0 L with deionized water giving a concentration of 4.1 g/L. The particles in this suspension did not precipitate. The stock suspension was administered to the test unit with a syringe pump. The flow rate of the syringe pump was 9.6 mL/day giving a nominal concentration of the test substance in the influent of the unit of 28.2 mg/L at a sewage supply rate of 1.4 L/day.

The CAS test was performed according to the study plan. The study plan was developed from ISO (1995), EC (1988) and OECD (1981) test guidelines. The test and control unit were not coupled. The units were started with activated sludge. The aeration was achieved by operating an air-lift.
The aeration rate was regulated so that the activated sludge was kept in suspension and the dissolved oxygen concentration was at least 2 mg/L. This oxygen concentration in the aeration vessel was measured at least two times a week. The domestic sewage supply was supplied at a rate of approximately 1.4 L/day to give a hydraulic retention time of 6 hours. The flow was checked by measuring the total volume of effluent over a 24-hour period.
After brushing, 35 mL of sludge was daily removed from the aeration tank to maintain a sludge retention time of 10 days. The effluent samples (50 mL) were taken from the settler. The NPOC values were primarily used to assess the performance of biological treatment system fed with ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives containing wastewater and to preliminary follow the removal of the test substance during the test period.
Specific analyses of ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-,N-(hydrogenated tallow alkyl) derivatives were used to determine the primary removal of the test substance.

Test performance:

The incubation temperature of both CAS units ranged from 19 to 21°C. The pH of the effluent of the CAS units varied from 7.0 to 7.3. The oxygen concentrations measured in both units were ≥ 2.9 mg/L . These test conditions are believed to allow biodegradation by micro-organisms present in activated sludge. The CAS test was started with a high concentration of aerobic micro-organisms (3.0 g/L dry weight) maintained by the daily addition of primary settled sewage. The daily removal of 35 mL of activated sludge from the aeration vessel resulted in a sludge retention time of 10 days. The dry weight in the CAS units ranged from 2.5 to 3.0 g/L . The performance of the control unit was checked by measuring the COD removal at Day 14 and at day 63 (day before prolongation of the test) and the concentrations of ammonium and nitrite in the effluent (Day 14). At Day 14 the COD contents in the influent and effluent were 420 and 43 mg/L, respectively. At day 63 the COD levels in the influent and effluent were 447 and 38 mg/L, respectively. COD removal percentages at were 90 (day 14) and 85 (day 62). The nitrite concentrations in the influent and effluent at Day 14 were <2.0 mg/L. The ammonium concentrations at Day 14 were 52 (influent) and <2.5 mg/L (effluent).
These results demonstrate that the test is valid

% Degr.:

99

St. dev.:

2

Parameter:

DOC removal

Sampling time:

47 d

Remarks on result:

other: 15 measurments obtained fromd day 47 to 62

% Degr.:

99.97

Parameter:

test mat. analysis

Sampling time:

61 d

Remarks on result:

other: (total removal from influent on 5 days from day 58 to 62) using tri (2-hydroxyethyl) octadecyl diamine as representative component

% Degr.:

3.8

Parameter:

test mat. analysis

Sampling time:

61 d

Remarks on result:

other: (removal from influent through adsorption onto sludge assessed in two samples day 61 and 62)

Transformation products:

no

Details on transformation products:

Complete mineralisation based on the organic carbon removal

Evaporation of parent compound:

no

Volatile metabolites:

no

Residues:

no

Details on results:

After the introduction of the test substance at day 0, high removal percentages of organic carbon (NPOC) were immediately accomplished.
These high removal percentages have to be ascribed to the adsorption and/or biodegradation of ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N- (hydrogenated tallow alkyl) derivatives. The removal percentages obtained during the first weeks of the test (62 to 96%) indicate formation of (a) water-soluble biodegradation product(s). After approximately 5 weeks biodegradation percentages of approximately 100 were achieved demonstrating that microorganisms in the CAS unit were capable of degrading the water-soluble biodegradation product(s). From Day 47 to 62 (end of test), samples were taken to assess a mean of the removal percentage with NPOC contents. No outliers were identified during day 47 to 62 using the Dixon test. Subsequently, the data obtained from Day 47 to 62 were used in a t-statistic. The mean difference between the NPOC in the effluents of control and test unit was 0.2 ± 0.4 mg/L (95 per cent confidence interval). The mean removal percentage calculated with this mean difference was 99 ± 2 (95% confidence). The t-statistic (n = 15) did not exceed the critical value and the mean difference is therefore statistically not significant. The results demonstrate that the continuous activated sludge system treating domestic wastewater spiked with ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives removes 100% of the organic carbon of the test substance.
The results indicate that ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives are degraded completely by microorganisms. The removal percentages demonstrate that formation of recalcitrant water-soluble substances is unlikely during the biodegradation process.

The concentration of ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives in the effluent of the test unit from day 58 to 62 using ethanol, 2,’2((3-((2-hydroxyethyl)amino)propyl)imino)bis-, octadecyl as representative component ranged from 4.4 to 8.5 µg/L. These concentrations correspond with >99,97% removal of the test substance from the influent. The concentrations of ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives in the mixed liquid suspended solids (activated sludge) of the reactor monitored on days 61 and 62 were 39.4 and 46.3 mg/L, respectively. Based on the observed mean test substance concentration in the mixed liquid
suspended solids of 42.9 mg/L. 3.8% removal of ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives by adsorption was calculated (Table IV). The results demonstrate that ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl)derivatives biodegrade almost completely in properly operating conventional biological wastewater treatment plants.

Validity criteria fulfilled:

yes

Remarks:

see test performance

Conclusions:

Study performed under GLP according guidelines with acceptable deviations. Meeting validity and quality criteria.
In conclusion, the CAS test demonstrates that ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives, are almost completely removed from the wastewater in conventional biological wastewater treatment plants. Ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives are primarily removed by biodegradation.

Executive summary:

The continuous activated sludge (CAS) test was performed according to ISO Guidelines, and in compliance with the OECD principles of Good Laboratory Practice. Ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N- (hydrogenated tallow alkyl) derivatives were exposed to micro-organisms maintained by addition of domestic wastewater in the CAS test. Ethanol, 2,2’-[[3- [(2-hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives were spiked at a nominal influent concentration of 28.2 mg/L (20.0 mg/L carbon; calculated) for a period of 62 days and included a control fed with domestic wastewater only. The immediate organic carbon removal percentages can be attributed to adsorption and probably biodegradation. The mean organic carbon removal percentage of ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N- (hydrogenated tallow alkyl) derivatives calculated over 15 measurements obtained from day 47 to 62 of the test was 99 +/-2% (95% confidence interval). The difference in organic carbon concentrations of the control and test measured is not statistically significant. This high removal percentage strongly indicates that ethanol, 2,2’-[[3-[(2-hydroxyethyl)amino]propyl]imino]bis-, N- (hydrogenated tallow alkyl) derivatives are biodegraded ultimately. Formation of water-soluble compounds during biological treatment of ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives can be excluded. An accurate assessment of the removal of ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives was established with specific analyses. The method (LC-MS/MS) for the determination of the test substance was satisfactory with regard to the linearity, repeatability of the injections, limit of quantification (LOQ), recovery, and specificity. The mean removal percentage of ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives in the test unit as quantified with the specific analysis from day 58 to 62 was >99.97%. These analyses demonstrate that the removal of ethanol, 2,2’-[[3 -[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives from the wastewater is complete. Ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives concentrations in the sludge of the reactor sampled on days 61 and 62 were 39.4 and 46.3 mg/L. The mean removal percentage of ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives from the influent through adsorption onto sludge assessed is therefore only 3.8%. The low removal percentage by adsorption demonstrates that ethanol, 2,2’-[[3- [(2-hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives are primarily removed by biodegradation. In conclusion, the CAS test demonstrates that ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives, are almost completely removed from the wastewater in conventional biological wastewater treatment plants. Ethanol, 2,2’-[[3-[(2- hydroxyethyl)amino]propyl]imino]bis-, N-(hydrogenated tallow alkyl) derivatives are primarily removed by biodegradation.

Reference 2

Endpoint:

biodegradation in water: sewage treatment simulation testing

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

28-10-2009 - 05-02-2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

See section 13.2 for the read-across justification.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 303 A (Simulation Test - Aerobic Sewage Treatment. A: Activated Sludge Units)

Deviations:

yes

Remarks:

minor acceptable deviations

Principles of method if other than guideline:

A few minor deviations to the guidelines were introduced. The primary settled sewage was collected weekly and stored in the refrigerator until
required instead of a daily collection of wastewater. The units consisted of aeration vessels capable of holding only 0.35 L from which the liquor
was then passed continuously to settler of 0.30 liter capacities

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

no

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

Secondary activated sludge to inoculate the test at the start was collected on 28-10-2009 from the wastewater treatment plant (WWTP) Nieuwgraaf in Duiven, The Netherlands. The WWTP Nieuwgraaf is an activated sludge plant treating predominantly domestic sewage. 0.35 liter of secondary activated sludge containing approximately 3 g/L dry weight was used as an inoculum for each CAS unit. This dry weight was obtained by diluting the sludge
obtained from the treatment plant. The primary settled sewage was collected from the same plant weekly and stored frozen until required.

Duration of test (contact time):

48 d

Initial conc.:

50 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

DOC removal

test mat. analysis

Details on study design:

CAS unit
The CAS test was performed in Hussmann-type units constructed of glass . The units consisted of an aeration vessel capable of holding 0.35 liter from which the liquor was passed continuously to a settler of 0.3 liter. The domestic waste water liquor in a cooled vessel was supplied with a pump. The liquor passed through the aeration vessel and settler and treated effluent left the apparatus to be collected in a vessel. Aeration was achieved through a capillary on the bottom of the aeration section at a rate of approximately 8 L/h of air. Sludge accumulating around the top of the aeration vessel was returned in the system once a day by brushing.

Stock suspension
A suspension of oleyl bis(2-hydroxyethyl)amine of 7.3 g/L in deionized water was directly added to the test unit using a syringe pump. The stock was prepared by adding 7.3 g of test substance to 0.7 L of deionized water. A homogenous suspension was obtained by acidifying the stock to a pH of
approximately 5.5 by adding HCl. A suspension was obtained by stirring for a few hours on a magnetic stirrer. The final stock suspension was made
up to 1.0 L with deionized water giving a concentration of 7.3 g/L. The particles in this suspension did not precipitate. The flow rate of the syringe
pump was 9.6 mL/day giving a nominal concentration of the test substance in the influent of the unit of 50 mg/L at a sewage supply rate of 1.4 L/day

Procedures of the CAS test
The CAS test was performed according to ISO (1995), EC (1988) and OECD (1981) test guidelines. The test and control unit were not coupled.
The units were started with activated sludge. The aeration was achieved by operating an air-lift. The aeration rate was regulated so that the activated
sludge was kept in suspension and the dissolved oxygen concentration was at least 2 mg/L. This oxygen concentration in the aeration vessel was
measured at least two times a week. The domestic sewage supply was supplied at a rate of approximately 1.4 L/day to give a hydraulic retention time
of 6 hours. The flow was checked by measuring the total volume of effluent over a 24-hour period. After brushing, 35 mL of sludge was daily
removed from the aeration tank to maintain a sludge retention time of 10 days. The effluent samples (50 mL) were taken from the settler.
The NPOC values were primarily used to assess the performance of biological treatment system fed with oleyl bis(2-hydroxyethyl)amine containing
wastewater and to preliminary follow the removal of the test substance during the test period.
NPOC values of the last period of the test were used to calculate the mean removal percentage. The daily removal percentages were calculated by the
following equation: 100 x (CT-(Ct-Cc)) / CT. Where CT is the carbon of the test compound measured as NPOC added to the settled sewage,
Ct is the carbon found as NPOC in the effluent of the CAS unit spiked with the test substance and Cc is the carbon found as NPOC in the effluent of
the control CAS unit.
The analysis values in the test and control unit were treated as paired observations. Outliers of the mean difference (Xd) series were eliminated
according to the Dixon test at a 95% probability level. From the set of 'n' paired observations the mean difference (Xd) and the standard deviation (Sd) were calculated. The Sd is calculated with the following formula (see attached report). The statistical significance of the observed difference was then
assessed from the t-statistics given by the following equation: (see attached report). The critical value of t at the required confidence level was\
obtained from statistical tables for a one tailed test with n-1 degrees of freedom. The percentage biodegradation/removal was given by;
(SL-Xd)/SL x 100 where Xd the mean difference and SL is the spiking level, both values being expressed in mg/L carbon.
The 95% confidence interval was calculated as follows: tn x Sd /SQRT(n) where tn is the t statistic for a two-tailed test, n-1 degrees of freedom,
P = 0.05.

Specific analyses of oleyl bis(2-hydroxyethyl)amine were used to determine the primary removal of the test substance. The removal percentage of
oleyl bis(2-hydroxyethyl)amine was determined with the following equation; (Is-Es)/Is x 100, where Is is the nominal test substance concentration in
the influent and Es is the mean of the measured test substance concentrations in the effluent.
The concentration of the test substance in the mixed liquid suspended solids (adsorbed on the activated sludge) (Csludge) and the theoretical
maximum concentration on sludge are used to assess the removal of the test substance by adsorption. Provided biodegradation nor evaporation of
the test substance occurs in the system, the theoretical maximum concentration of oleyl bis(2-hydroxyethyl)amine adsorbed onto the sludge is;
Cmax adsorption = Is x SRT/HRT, where SRT is the sludge retention time, HRT is the hydraulic retention time (both expressed in days) and Is is the
nominal test substance concentration in the influent. The removal of oleyl bis(2-hydroxyethyl)amine by adsorption is;
removal (%) = 100 x Csludge/Cmax adsorption.

Reference substance:

not required

Test performance:

Test conditions and validity of the test
The incubation temperature of both CAS units ranged from 19 to 21°C. The pH of the effluent of both CAS units varied from 7.0 to 7.4.
The oxygen concentrations measured in both units were always ≥3.9 mg/L (Table I). These test conditions are believed to allow biodegradation by
micro-organisms present in activated sludge.
The CAS test was started with a high concentration of aerobic micro-organisms (3.0 g/L dry weight) maintained by the daily addition of primary
settled sewage and sludge from a full-scale treatment plant. The daily removal of 35 mL of activated sludge from the aeration vessel resulted in a
sludge retention time of 10 days. The dry weight in the CAS units ranged from 2.4 to 3.0 g/L (Table I).
The performance of the control unit was checked by measuring the COD removal at Day 14 and at day 48 and the concentrations of ammonium and
nitrite in the effluent (Day 14). At Day 14 the COD contents (mean of two measurements) in the influent and effluent were 416 and 43 mg/L,
respectively. At day 48, the COD levels in the influent and effluent were 461 and 38 mg/L, respectively. COD removal percentages at both days were
90 and 92. The ammonium and nitrite concentrations in the effluent at Day 14 were <2.5 and <2.0 mg/L. These results demonstrate that the test is valid.

% Degr.:

ca. 100

St. dev.:

0.8

Parameter:

DOC removal

Remarks on result:

other: 15 measurements from day 34 - 48

% Degr.:

> 99.99

Parameter:

test mat. analysis

Remarks on result:

other: analysed in effleunt of test unit from day 44 to 48

% Degr.:

0.16

Parameter:

test mat. analysis

Remarks on result:

other: removal from influent throug adsorption onto sludge assessed in two samples day 47 and 48

Transformation products:

no

Details on transformation products:

These high NPOC removal percentages strongly indicate that oleyl bis(2-hydroxyethyl)amine is biodegraded completely. Formation of water soluble
compounds during biological treatment of oleyl bis(2-hydroxyethyl)amine can be excluded.

Evaporation of parent compound:

no

Volatile metabolites:

no

Residues:

no

NPOC concentrations in the effluent of the control and test unit and the calculated removal percentages of oleyl bis(2-hydroxyethyl)amine. The data in grey part of the table are used to calculate the biodegradation percentage.

Time (days)	NPOC (mg/L)		Removal (%)
	Control	Test
-4	13.7	11.4
-2	12.1	10.9
2	9.5	11.4	95
0	10.8	18.2	80
6	11.0	11.3	99
9	11.7	10.7	103
13	12.7	10.2	107
16	13.9	16.2	94
20	12.7	12.1	102
23	8.3	8.8	99
27	9.7	11.8	94
30	7.5	9.4	95
34	9.1	8.2	102
35	12.0	9.5	107
36	10.3	8.0	106
37	12.7	9.3	109
38	10.2	8.8	104
39	10.9	8.2	107
40	11.0	11.3	99
41	10.5	10.2	101
42	11.4	10.5	102
43	13.0	14.9	95
44	12.1	12.2	100
45	13.9	13.7	101
46	12.8	12.3	101
47	12.6	12.5	100
48	13.7	13.9	100

Concentrations ofoleyl bis(2-hydroxyethyl)aminemeasured in the effluent and mixed liquid suspended solids (adsorption onto sludge) and removal percentages from the influent and by adsorption onto sludge, respectively.

Time (days)	Concentration (μg/L)	Removal (%)
	Effluent
44	<0.06	>99.999
45	<0.06	>99.999
46	<0.06	>99.999
47	<0.06	>99.999
48	<0.06	>99.999
Time (days)	Concentration (mg/L)	Removal (%)
	Mixed liquid suspended solids
47	3.2	0.16
48	3.2	0.16

Validity criteria fulfilled:

yes

Remarks:

COD removal in the control unit at Day 14 and day 48 of 90 and 92% resp. The ammonium and nitrite concentrations in the control effluent at Day 14 were <2.5 and <2.0 mg/L. These results demonstrate that the test is valid

Conclusions:

The CAS test demonstrates that oleyl bis(2-hydroxyethyl)amine almost completely removed from the wastewater in conventional biological
wastewater treatment plants. Oleyl bis(2-hydroxyethyl)amine is primarily removed by biodegradation.
102±0.8% NPOC removal, determined in effluent samles from test unit during 15 days from day 34 to 48
>99.999% removal of test substance calculated with specific analysis in effluent samples taken from day 44-48
0.16 % removal of test substance through adsorption onto sludge calculated with specific analysis in sludge samples taken on day 47 and 48.

Test performed under GLP according guidelines with a few accpetable (minor) deviations, meeting all validity criteria

Executive summary:

The continuous activated sludge (CAS) test was performed according to ISO Guidelines, and in compliance with the OECD principles of Good Laboratory Practice.Oleyl bis(2-hydroxyethyl)amine was exposed to micro-organisms maintained by addition of domestic wastewater in the CAS test.

Oleyl bis(2-hydroxyethyl)amine was spiked at a nominal influent concentration of 50 mg/L (37.0 mg/L carbon; calculated) for a period of 48 days and included a control fed with domestic wastewater only.

The immediate high removal percentages can be attributed to adsorption and probably biodegradation. The mean removal percentage of oleyl bis(2-hydroxyethyl)amine calculated over 15 measurements obtained from day 34 to 48 of the test was 102±0.8% (95% confidence interval). These high removal percentages strongly indicate that oleyl bis(2-hydroxyethyl)amine is biodegraded completely. Formation of water soluble compounds during biological treatment ofoleyl bis(2-hydroxyethyl)amine can be excluded.

An accurate assessment of the removal of oleyl bis(2-hydroxyethyl)amine was established with specific analyses. The method (LC-MS/MS) for the determination of oleyl bis(2-hydroxyethyl)amine was valid with regard to the linearity, repeatability of the injections, limit of quantification (LOQ), limit of detection (LOD), recovery and system stability. The mean removal percentage of oleyl bis(2-hydroxyethyl)amine in the test unit was quantified with the specific analysis from day 44 to 48 was >99.999% using octadecenyl bis(2-hydroxyethyl) amine as representative component as a worst-case. These analyses demonstrate that the removal of oleyl bis(2-hydroxyethyl)amine is complete. Oleyl bis(2-hydroxyethyl)amine concentrations in the mixed liquid suspended solids (activated sludge) of the reactor sampled on days 47 and 48 were 3.2 mg/L. Mean removal percentages of oleyl bis(2-hydroxyethyl)amine reaction from the influent through adsorption onto sludge assessed in two samples was therefore 0.16 % demonstrating that oleyl bis(2-hydroxyethyl)amine is primarily removed by biodegradation.

In conclusion, the CAS test demonstrates that oleyl bis(2-hydroxyethyl)amine almost completely removed from the wastewater in conventional biological wastewater treatment plants.Oleyl bis(2-hydroxyethyl)amineis primarily removed by biodegradation.

Description of key information

Sewage Treatment Plant

No sewage treatment simulation tests for degradation rates under environmental conditions are available for primary fatty amine propoxylates but for two very similar substances di (2-hydroxyethyl) oleylamine (PFAEO O, CAS 25307-17-9)  and tri (2-hydroxyethyl) hydrogenated tallow diamine (PFAEO HT, CAS 1218787-30-4, old CAS 90367-25-2) STP simulation test results are available and these results will be used for read-across to primary fatty amine propoxylates (PFAPO-T).

A removal percentage of >99.97 is adopted as a realistic worst-case removal in an STP for di (2-hydroxypropyl) tallow amine based on the removal determined in a wastewater simulation test with tri (2-hydroxyethyl) hydrogenated tallow diamine. The adopted removal of di (2-hydroxypropyl) tallow amine through adsorption in a biological treatment plant of 1.63% is based on linear interpolation of the removal percentages of di (2- hydroxyl ethyl) oleyl amine and tri (2-hydroxyethyl) hydrogenated tallow diamine due to sorption when related to the calculated Koc.

Sediment

No half-life data are available for sediment. The half-life as observed for soil of 90 days is therefore used for aerobic sediment similar as in EUSES as there is no principal difference between soil and sediments on respect to the sorption properties, as a worst case approach the value for soil is also used for aerobic sediment.

Key value for chemical safety assessment

Half-life in freshwater:

42 d

at the temperature of:

12 °C

Half-life in freshwater sediment:

90 d

at the temperature of:

12 °C

Additional information

Removal in biological waste water treatment systems

Behavior of organic substances in biological wastewater treatment systems is mainly determined by their biodegradability, capacity to adsorb, and evaporation. SimpleTreat does integrate adsorption, stripping and biodegradation (Struijs et al., 1991, Struijs, 1996). The justness of read-across of removal of organic substances from wastewater in biological treatment systems should therefore be based on these properties. The fate of di (2 -hydroxypropyl) tallow amine (PFAPO T) in wastewater treatment systems is assessed through read across with di (2-hydroxy ethyl) oleyl amine (PFAEO O). The removal of di (2-hydroxy ethyl) oleyl amine and tri (2-hydroxy ethyl) hydrogenated tallow diamine (as supporting study) have been determined in CAS tests.

The removal of the readily biodegradable di (2-hydroxyl ethyl) oleyl amine (CAS no: 25307-17-9) and tri (2-hydroxy ethyl) hydrogenated tallow diamine (CAS no 90367-25-2) in biological treatment plants was assessed in continuously-fed activated sludge (CAS) unit fed with domestic wastewater spiked with the respective test substances (Akzo Nobel, 2010 and 2011). Di (2-hydroxyl ethyl) oleyl amine and tri (2-hydroxyethyl) hydrogenated tallow diamine (as supporting study) were exposed to micro-organisms maintained by addition of domestic wastewater in the CAS test. Both substances were spiked at nominal influent concentrations of 50 mg/L (resp. 37.0 and 28.2 mg/L carbon; calculated) for a period of resp. 48 and 62 days, respectively, and included a control fed with domestic wastewater only.

The mean carbon removal percentage of di (2- hydroxyl ethyl) oleyl amine calculated over 15 measurements of the test was 102± 0.8% (95% confidence interval). These high removal percentages strongly indicate that di (2-hydroxyl ethyl) oleyl amine is biodegraded completely. Formation of water soluble compounds during biological treatment of di (2-hydroxyl ethyl) oleyl amine can be excluded. An accurate assessment of the removal of di (2-hydroxyl ethyl) oleyl amine was established with specific analyses. The mean removal percentage of di (2-hydroxyl ethyl) oleyl amine in the test unit was >99.999% using octadecenyl bis(2-hydroxyethyl) amine as most representative component. These analyses demonstrate that the removal of di (2-hydroxyl ethyl) oleyl amine is complete. Mean removal percentages of octadecenyl bis(2-hydroxyethyl) amine from the influent through adsorption onto sludge assessed in two samples was 0.16% demonstrating that octadecenyl bis(2-hydroxyethyl) amine is primarily removed by biodegradation.

The mean carbon removal percentage of tri (2-hydroxyethyl) hydrogenated tallow diamine calculated over 15 measurements of the test was 99 ± 2% (95% confidence interval). These high removal percentages strongly indicate that tri (2-hydroxyethyl) hydrogenated tallow diamine is biodegraded completely. Formation of water soluble compounds during biological treatment of tri (2-hydroxyethyl) hydrogenated tallow diamine can be excluded. An accurate assessment of the removal of tri (2-hydroxyethyl) hydrogenated tallow diamine was established with specific analyses. The mean removal percentage of tri (2-hydroxyethyl) hydrogenated tallow diamine in the test unit was >99.97.% using tri (2-hydroxyethyl) octadecyl diamine as most representative component. Mean removal percentages of tri (2-hydroxyethyl) hydrogenated tallow diamine from the influent through adsorption onto sludge assessed in two samples was 3.8% demonstrating that tri (2-hydroxyethyl) hydrogenated tallow diamine is primarily removed by biodegradation.

 

More than 99.999% removal is observed for 2,2’(octadec-9-enylimino)-bisethanol (25307-17-9) in waste water treatment simulation test. From this removal 99.83% is removed via biodegradation and 0.16 % via sorption. The test result can be used for the other primary fatty amine ethoxylates as a worst-case because all primary fatty amine ethoxylates

The removal ofoctadecenyl bis(2-hydroxyethyl)amine in biological treatment plants was simulated in a continuously-fed activated sludge (CAS) unit fed with domestic wastewater spiked with octadecenyl bis(2-hydroxyethyl)amine (AkzoNobel, 2010). Octadecenyl bis(2-hydroxyethyl)amine was exposed to micro-organisms maintained by addition of domestic wastewater in the CAS test. Octadecenyl bis(2-hydroxyethyl)amine was spiked at a nominal influent concentration of 50 mg/L (37.0 mg/L carbon; calculated) for a period of 48 days and included a control fed with domestic wastewater only.

The immediate high removal percentages can be attributed to adsorption and probably biodegradation. The mean removal percentage of octadecenyl bis(2-hydroxyethyl)amine calculated over 15 measurements obtained from day 34 to 48 of the test was 102±0.8% (95% confidence interval). These high removal percentages strongly indicate that octadecenyl bis(2-hydroxyethyl)amine is biodegraded completely. Formation of water soluble compounds during biological treatment of octadecenyl bis(2-hydroxyethyl)amine can be excluded. An accurate assessment of the removal of octadecenyl bis(2-hydroxyethyl)amine was established with specific analyses. The mean removal percentage of octadecenyl bis(2-hydroxyethyl)amine in the test unit was quantified from day 44 to 48 and was more than 99.999% usingoctadecenyl bis(2-hydroxyethyl) amineas representative component as a worst-case. These analyses demonstrate that the removal of octadecenyl bis(2-hydroxyethyl)amine is complete. Octadecenyl bis(2-hydroxyethyl)amine concentrations in the mixed liquid suspended solids (activated sludge) of the reactor sampled on days 47 and 48 were 3.2 mg/L. Mean removal percentages of octadecenyl bis(2-hydroxyethyl)amine from the influent through adsorption onto sludge assessed in two samples was therefore 0.16% demonstrating that octadecenyl bis(2-hydroxyethyl)amine is primarily removed by biodegradation.

In conclusion, the CAS test demonstrates thatoctadecenyl bis(2-hydroxyethyl)amine is almost completely removed from the wastewater in conventional biological wastewater treatment plants.Octadecenyl bis(2-hydroxyethyl)amineis primarily removed by biodegradation.

 

In conclusion, the CAS test demonstrates that both ethoxylated fatty amine derivatives are almost completely removed from the wastewater in conventional biological wastewater treatment plants and this is also assumed for Di (2-hydroxypropyl) tallow amine (PFAPO T). These type of surfactants are primarily removed by biodegradation. A further explanation can also be found within the read-across justification in chapter 13.

 

 

The biodegradability, capacity to adsorb and evaporation of the three alkoxylated fatty amines are given in the Table below. The alkoxylated fatty amine derivatives have been classified as readily biodegradable. The Henry’s Law constants of the surfactants demonstrate that evaporation is negligible. The Log Koc indicates that the potential of di (2-hydroxypropyl) tallow amine to adsorb onto activated sludge is in between of those of di (2- hydroxy ethyl) oleyl amine and tri (2-hydroxy ethyl) hydrogenated tallow diamine.The properties of di (2- hydroxyl ethyl) oleyl amine (CAS no: 25307-17-9), di (2-hydroxypropyl) tallow amine (Old CAS no: 68951-72-4, new CAS 1309955-79-0) and tri (2-hydroxyethyl) hydrogenated tallow diamine(CAS no 90367-25-2) demonstrate that read across is justified. A further read-across justification is included in chapter 13.

A removal percentage of >99.97 is adopted as a worst-case for di (2-hydroxypropyl) tallow amine based on the removal determined in a wastewater simulation test with tri (2-hydroxyethyl) hydrogenated tallow diamine. The adopted removal of di (2-hydroxypropyl) tallow amine through adsorption in a biological treatment plant of 1.63% is based on linear interpolation of the removal percentages of di (2- hydroxyl ethyl) oleyl amine and tri (2 -hydroxyethyl) hydrogenated tallow diamine due to sorption when related to the calculated Koc.

 

 

Table: Biodegradability test results (OECD 301 D), Henry’s law constant and the Log Koc of the ethoxylated/propoxylated fatty amine derivatives and removal percentages of di (2- hydroxy ethyl) oleyl amine and tri (2-hydroxy ethyl) hydrogenated tallow diamine determined in CAS tests (OECD 303). The removal percentages of di (2-hydroxypropyl) tallow amine obtain trough read across are indicated in red.

Substance	Biodegradability	Henry’s Law constant Pa.m3/mol (Evaporation potential*)	Log Koc L/kg (Potential to adsorb)	Total removal in CAS test %	Removal via sorption in CAS test %
Di (2- hydroxy ethyl) oleyl amine (PFAEO O) (CAS no: 25307-17-9)	Readily	1.215 * 10 ^-5(calculated EPI)   0.071 (calculated from ratio measured VP/CMC)   (Negligible)	Log Koc = 3.70 (calculated, EPI)   Log Koc = 4.96 (measured, range 4.61 – 4.97)	>99.999 (measured)	0.16 (measured)
Di (2-hydroxy propyl) tallow amine (PFAPO T) (Old CAS no: 68951-72-4, New CAS no: 1309955-79-0)	Readily biodegradable and showing ultimate biodegradatin in SCAS test	1.885 * 10 ^-4(calculated EPI)   2.2 (calculated from ratio VP/S)   (Negligible)	Log Koc = 4.19 (calculated, EPI) Read across from Bis (2 -hydroxyethyl) oleyl amine (PFAEO O)	>99.97 ( worst-case)	1.63 (interpolation in relation to calculated Koc)
Tri (2-hydroxy ethyl) hydrogenated tallow diamine (CAS no: 90367-25-2)	Readily	2.878 * 10^-10(calculated EPI)     (Negligible)	Log Koc = 4.91 (calculated, EPI) Log Koc = 5.81 (measured, range 5.09 – 6.26)	>99.97 (measured)	3.8 (measured)

*If the Henry‟s law constant is in the range 10^-2< kH < 1 Pam³/ mol, the test substance volatilizes slowly at a rate dependent on the Henry‟s law constant. If the Henry’s law constant is in the range 1 < kH < 10²Pam³/mol, the test substance volatilizes not rapid but possibly significant.

 

Biodegradation in sediment:

Default half-life’s

For the derivation of the default half-life’s the bioavailability is taken into account via the sorption properties of the substance. This is realistic for soluble (non)-toxic substances. For poorly soluble/strongly sorbing substances however, the degradation rate in a standard ready test is limited by the dissolution rate and in many cases not 60% degradation is achieved within 28 days but in a slightly longer time frame. Such a substance is in fact completely degraded to CO₂and H₂O and thus completely biodegradable. The bioavailable fraction is readily biodegradable but due to the stringency of the test setup the substance this cannot be observed.

The Kpsoil was determined as 4526 L/kg for a highly similar substance (PFAEO O, CAS 25307-17-9). The suggested maximum half-life for a readily degradable substance with a Kpsoil in the range >1000 and < 10000 L/kg is 3000 days for soil (at 12 ºC). These values are considered as extremely conservative but in the absence of measured data can be used in the exposure assessment as a worst-case. The half-life of the bioavailable fraction of primary fatty amine ethoxylates and propoxylates in the water phase of soils is expected to be in the order of a few days, which is based on experiments with dialkyldimethylammonium salts (van Ginkel et al, 2003).

Read across from primary alkyl amines 

No half-life data are available for sediment. The half-life as observed for soil of 90 days is therefore used for aerobic suspended sediment similar as in EUSES as there is no principal difference between soil and sediments on respect to the sorption properties, as a worst-case approach, the value for soil is also used for aerobic sediment.