Reaction product of p-nonylphenol, formaldehyde and diethanolamine, propoxylated

Biodegradation in water and sediment: simulation tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: simulation testing on ultimate degradation in surface water

Type of information:

experimental study

Adequacy of study:

key study

Study period:

06 May 2020 to 18 Sep 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 309 (Aerobic Mineralisation in Surface Water - Simulation Biodegradation Test)

Version / remarks:

2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

no

Oxygen conditions:

aerobic

Inoculum or test system:

natural water

Details on source and properties of surface water:

An overview of the Sampling Site, Sampling Conditions and Water Characteristics is provided in Table 1 in 'Any other information on materials and methods incl. tables'.
- Details on collection: The water was collected from the surface of the river 'Leine' (approx. upper 20 cm water) at an undisturbed recess. Furthermore, temperature, pH value and O2 concentration of the water were measured at field sampling.
- Location of collection: Am Anger, 30982 Pattensen, Germany
- Temperature at time of collection: 12.2 °C
- pH at time of collection: 7.64
- Oxygen concentration: 9.43 mg O2/L
- Dissolved organic carbon: 5.21 mg/L
- Handeling: The water was clear and colourless. Small fishes were removed by filtration.
- Suspended mater: To minimize possible interferences of NER formation with biodegradation and to allow proper kinetic evaluation, water with an amount of suspended matter < 10 mg/L was used.
- Acclimation: Until test start the surface water was acclimated under test conditions at 12 ± 2 °C in the dark with aeration.
- Storage: Storage at 4 ± 2 °C was not necessary as the test was started within 2 days after collection of the water.

Details on source and properties of sediment:

Not applicable

Details on inoculum:

Not applicable

Duration of test (contact time):

90 d

Initial conc.:

20 µg/L

Based on:

test mat.

Initial conc.:

100 µg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

other: The amount of test item constituents and degradation products was determined

Details on study design:

TEST CONDITIONS
- Volume of test solution/treatment: 80 mL (test item replicates, sterile controls and sterile blank controls); 100 mL (blank controls); 150 mL (reference control and solvent control)
- Test temperature: 12 ± 2 °C
- pH: 7.50 – 7.85
- Oxygen concentration: 9.29 – 9.60 mg O2/L
- Agitation: Continuous stirring at approx. 120 rpm to maintain particles and microorganisms in suspension and to facilitate oxygen transfer from the headspace to the liquid.
- Continuous darkness: Yes

TEST SYSTEM
- Culturing apparatus: 120 mL gastight glass bottles for test item replicates, sterile controls and sterile blank controls. 150 mL gastight glass bottles for blank controls. 300 mL gastight bottles for reference control and solvent control replicates.
- Number of replicates for test item: 2/sample/ concentration (in total 28 replicates were prepared for each concentration)
- Number of replicates for sterile controls: 2/sample/ concentration (in total 16 replicates were prepared for each concentration)
- Number of replicates for blank controls: 1 (in total 10 replicates were prepared for each concentration)
- Number of replicates for sterile blank controls: 1 (in total duplicates were prepared)
- Number of replicates for reference control: 2
- Number of replicates for solvent control: 2

SAMPLING
- Temperature: The incubation temperature was recorded continuously throughout the test.
- pH and Oxygen: Measurements of pH and oxygen concentration in the test system (Blank control) were done on day 0, 33, 40, 62 and 89.
- Transformation: Sampling for determination of the transformation was carried out directly after application and at 13 additional sampling points. The sampling points were chosen to enable the establishment of the pattern of decline of the test item constituents. Sampling was done after application (day 0) and at day 2, 5, 8, 12, 14, 20, 33, 43, 51, 57, 62, 75 and day 90. For the reference control sampling was stopped once degradation was finished. Sampling was done at day 0, day 6 and day 12. Residual activity in the test solution and dissolved 14CO2 (after acidification of test solution) was determined by LSC.
- Sampling method used per analysis type: The amount of test item constituents and degradation products in the test solution and sterile test item solution was determined by LC-HRMS. Two replicates per test item concentration were harvested at each sampling time for determination of transformation until day 62. At day 75 and day 90 the remaining four replicates were analysed. Sterile controls were analysed in parallel at each sampling point. One blank control and one sterile blank control were analysed in parallel to the respective sampling.

Reference substance:

other: Aniline sulfate, [14C(U)]-

Remarks:

10 μg/L

Compartment:

natural water: freshwater

% Recovery:

92

Remarks on result:

other: Constituent 9; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

96

Remarks on result:

other: Constituent 10; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

90

Remarks on result:

other: Constituent 6; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

95

Remarks on result:

other: Constituent 1; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

94

Remarks on result:

other: Constituent 7; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

96

Remarks on result:

other: Constituent 2; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

88

Remarks on result:

other: Constituent 3; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

87

Remarks on result:

other: Constituent 4; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

89

Remarks on result:

other: Constituent 5; 20 µg/L

Compartment:

natural water: freshwater

% Recovery:

100

Remarks on result:

other: Constituent 9; 100 µg/L

Compartment:

natural water: freshwater

% Recovery:

100

Remarks on result:

other: Constituent 10; 100 µg/L

Compartment:

natural water: freshwater

% Recovery:

100

Remarks on result:

other: Constituent 6; 100 µg/L

Compartment:

natural water: freshwater

% Recovery:

85

Remarks on result:

other: Constituent 1; 100 µg/L

Compartment:

natural water: freshwater

% Recovery:

99

Remarks on result:

other: Constituent 7; 100 µg/L

Compartment:

natural water: freshwater

% Recovery:

100

Remarks on result:

other: Constituent 2; 100 µg/L

Compartment:

natural water: freshwater

% Recovery:

100

Remarks on result:

other: Constituent 3; 100 µg/L

Compartment:

natural water: freshwater

% Recovery:

100

Remarks on result:

other: Constituent 4; 100 µg/L

Compartment:

natural water: freshwater

% Recovery:

100

Remarks on result:

other: Constituent 5; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

100

Parameter:

test mat. analysis

Sampling time:

62 d

Remarks on result:

other: all studied constituent; 20 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

99

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 9; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

98

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 10; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

99

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 11; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

95.8

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 6; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

94

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 1; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

95.2

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 7; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

95.2

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 2; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

95.2

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 3; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

95.2

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 4; 100 µg/L

Parent/product:

parent

Compartment:

water

% Degr.:

95.2

Parameter:

test mat. analysis

Sampling time:

90 d

Remarks on result:

other: Constituent 5; 100 µg/L

Key result

Compartment:

natural water: freshwater

DT50:

25.2 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 5; 20 μg/L

Key result

Compartment:

natural water: freshwater

DT50:

37.3 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 1; 100 μg/L

Compartment:

natural water: freshwater

DT50:

19.6 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: Constituent 9; 20 μg/L

Compartment:

natural water: freshwater

DT50:

21.2 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: Constituent 10; 20 μg/L

Compartment:

natural water: freshwater

DT50:

19.4 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: Constituent 6; 20 μg/L

Compartment:

natural water: freshwater

DT50:

21.8 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 7; 20 μg/L

Compartment:

natural water: freshwater

DT50:

19.6 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 2; 20 μg/L

Compartment:

natural water: freshwater

DT50:

22.5 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 3; 20 μg/L

Compartment:

natural water: freshwater

DT50:

24 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 4; 20 μg/L

Compartment:

natural water: freshwater

DT50:

13.9 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 9; 100 μg/L

Compartment:

natural water: freshwater

DT50:

13.7 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 10; 100 μg/L

Compartment:

natural water: freshwater

DT50:

11.9 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 11; 100 μg/L

Compartment:

natural water: freshwater

DT50:

13.8 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 6; 100 μg/L

Compartment:

natural water: freshwater

DT50:

16.2 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 7; 100 μg/L

Compartment:

natural water: freshwater

DT50:

13.7 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 2; 100 μg/L

Compartment:

natural water: freshwater

DT50:

14.8 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 3; 100 μg/L

Compartment:

natural water: freshwater

DT50:

19.7 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 4; 100 μg/L

Compartment:

natural water: freshwater

DT50:

25.2 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: constituent 5; 100 μg/L

Transformation products:

not specified

Remarks:

M4, M5, M9, M10, M11, M20, M13, M14, M17, M18 and M19

Details on transformation products:

An overview of the transformation products is provided in Table 12 - Table 13 in ‘Any other information on results incl. tables’.
Metabolites were formed by oxidation of the primary or secondary alcohol moieties of the constituents. Eleven metabolites (M) were identified in the test. The link between metabolites and corresponding constituents of the test item is list below:
M4 and M5 were identified in Constituent 6 transformation products
M9 was identified in Constituent 7 transformation products
M10 and M11 were identified in Constituent 2 transformation products
M13, M14 and M20 were identified in Constituent 3 transformation products
M17, M18 and M19 were identified in Constituent 4 transformation products

The kinetic evaluations for the metabolites were performed based on the FOCUS guidance document on estimating persistence and degradation kinetics. DTx values were calculated based on single first order kinetics (SFO), the metabolite data were normalized for these calculations. The highest occurrence of a metabolite was fixed at 100 % correspondingly the respective day was set to day 0. The results indicate that the DT50 values of the metabolites at 100 µg/L applied concentration in surface water were in a range of 7.57 to 14.4 days. An overview of the results is provided in Table 14 in ‘Any other information on results incl. tables’.

Evaporation of parent compound:

not specified

Volatile metabolites:

no

Residues:

yes

Details on results:

An overview of the results is provided in Table 4 – Table 11 in ‘Any other information on materials and methods incl. tables’.
The test substance cannot be radiolabelled, because it is a UVCB substance. Due to the use of an unlabelled test item, analysis of Non-extractable residues (NER) was not possible.

Transformation of most constituents started in within 5 – 12 days after application. About 50% transformation was reached within 33 - 43 days, after 62 days the detected concentration was < LOQ or even not detectable. For constituent 1 transformation started only on day 43 and the transformation reached 50% after 57 days and 65 % on day 62. At day 75 the concentration was < LOQ.
In the sterile controls, no significant decrease was determined until day 90 for the constituents 9, 10 and 11. For constituent 1 - 7 no significant decrease was determined until day 43, thereafter the concentration decreased slightly until day 90.
The difference in dissipation (transformation) in the biotic test item replicates and the abiotic sterile controls clearly indicate that biodegradation is the major pathway of dissipation (transformation).

The kinetic evaluations were done based on the FOCUS guidance document on estimating persistence and degradation kinetics.
The kinetic models were chosen based on the following criteria:
• Visual assessment of the fitted and observed data versus time
• Visual assessment of the residuals
• Estimation of the error percentage at which the x2-test was passed

Results with reference substance:

An overview of the results is provided in Table 3 in ‘Any other information on results incl. tables’
Mineralisation of the reference item Aniline sulphate, [14C(U)]- in the reference control was 60% after 6 days and 65.6% after 12 days. In the solvent control the mineralisation was 50 % after 6 days and 64% after 12 days. The high mineralization indicates that the surface water contained an active microbial population and the microbial activity was not inhibited by the solvent.

Table 3 Mineralization of Aniline Sulphate, [14C(U)]- in the Reference Control and Solvent Control

Exposure Day	Reference Control		Solvent Control
	% Mineralisation1)		% Mineralisation1)
	Repl. 1	Repl. 1	Repl. 2	Repl. 2
6	59.7	60.3	50.8	49.4
12	66.0	65.2	64.0	64.5

1) calculated from residual activity in test solution (after acidification) and applied radioactivity

Table 4. Recovery Rates Procedural Recovery Samples: 20 µg/L (Approach 2)

Fortified concentrations (n = 1 per sampling day): 2 x LOQ: 15 µg/L

Sampling Day	Constituent 9	Constituent 10	Constituent 6	Constituent 1	Constituent 7	Constituent 2
	RR [%]	RR [%]	RR [%]	RR [%]	RR [%]	RR [%]
0	93	100	92	651)	90	95
2	84	87	86	81	83	92
5	96	110	97	88	96	96
8	81	93	87	112	82	97
12	95	107	100	107	103	96
14	101	89	95	85	95	88
20	91	88	96	88	97	97
33	95	107	113	146	122	107
43	86	91	86	79	84	90
51	85	92	89	85	87	94
57	88	96	95	108	96	91
62	92	96	90	95	94	96
Mean	84	89	87	87	87	88
Sampling Day	Constituent 3	Constituent 4	Constituent 5
	RR [%]	RR [%]	RR [%]
0	98	85	100
2	90	95	100
5	104	95	102
8	94	92	85
12	95	98	108
14	89	102	114
20	92	96	88
33	109	114	97
43	83	81	93
51	98	94	96
57	106	96	93
62	88	87	89
Mean	88	87	90

Table 5. Recovery Rates Procedural Recovery Samples: 100 µg/L (Approach 1)

Fortified concentrations (n = 1 per sampling day): 2 x LOQ: 15 µg/L

Sampling Day	Constituent 9	Constituent 10	Constituent 6	Constituent 1	Constituent 7	Constituent 2
	RR [%]	RR [%]	RR [%]	RR [%]	RR [%]	RR [%]
0	120	135	118	128	112	126
2	100	115	100	83	100	99
5	102	107	97	106	95	114
8	93	108	98	113	100	105
12	103	112	98	108	103	94
14	90	107	106	70	102	101
20	90	101	102	82	104	97
33	82	95	88	75	98	88
43	99	105	87	97	84	77
51	85	86	91	104	98	105
57	126	97	95	88	95	100
62	124	101	107	119	99	108
75	108	89	113	91	111	111
90	113	101	103	85	99	106
Mean	103	104	100	96	100	102
Sampling Day	Constituent 3	Constituent 4	Constituent 5
	RR [%]	RR [%]	RR [%]
0	126	136	123
2	100	105	113
5	108	105	91
8	103	102	118
12	102	99	93
14	108	124	107
20	104	101	103
33	92	82	88
43	80	81	71
51	100	109	97
57	96	97	102
62	108	101	108
75	111	113	129
90	114	106	115
Mean	104	104	104

Table 6. 20 µg/L test item

		% Applied (nominal)
Constituent	Repl.	Day
		0	2	5	8	12	14
Constituent 9	1	112.1	115.6	113.3	96.2	108.0	85.9
	2	104.6	112.2	105.4	104.4	85.1	58.1
	Mean	108.4	113.9	109.3	100.3	96.6	72.0
Constituent 10	1	105.2	99.9	118.1	84.7	90.3	76.2
	2	124.1	112.0	117.3	113.3	92.1	64.1
	Mean	114.7	105.9	117.7	99.0	91.2	70.1
Constituent 6	1	87.5	77.1	71.3	62.5	68.2	57.9
	2	90.3	77.7	77.9	73.2	62.4	48.8
	Mean	88.9	77.4	74.6	67.9	65.3	53.3
Constituent 1	1	126.1	90.7	181.1	111.7	108.4	101.0
	2	131.6	121.5	176.6	125.9	82.2	91.2
	Mean	128.9	106.1	178.8	118.8	95.3	96.1
Constituent 7	1	85.3	73.1	67.9	56.8	70.6	63.2
	2	92.8	73.0	87.1	75.7	57.5	50.7
	Mean	89.1	73.1	77.5	66.3	64.1	57.0
Constituent 2	1	103.8	99.7	100.4	81.2	88.4	67.3
	2	108.0	97.2	100.5	78.9	65.9	55.1
	Mean	105.9	98.5	100.5	80.1	77.1	61.2
Constituent 3	1	95.6	93.9	91.0	74.9	76.0	75.4
	2	96.7	88.7	92.4	81.5	71.1	60.4
	Mean	96.2	91.3	91.7	78.2	73.6	67.9
Constituent 4	1	91.4	94.4	95.9	74.7	84.0	79.5
	2	98.3	95.0	99.2	78.6	72.8	80.1
	Mean	94.8	94.7	97.6	76.7	78.4	79.8
Constituent 5	1	98.9	100.8	87.5	74.3	87.3	88.6
	2	101.6	96.5	94.1	90.7	75.5	62.5
	Mean	100.2	98.7	90.8	82.5	81.4	75.5

 

Table 6 (continued). 20 µg/L test item

		% Applied (nominal)
Constituent	Repl.	Day
		20	33	43	51	57	62
Constituent 9	1	62.2	23.8	<LCL	<LCL	<LCL	<LCL
	2	54.7	21.3	<LCL	<LCL	<LCL	n.d.
	Mean	58.5	22.6	<LCL	<LCL	<LCL	<LCL
Constituent 10	1	65.0	<LCL	n.d.	n.d.	n.d.	n.d.
	2	53.8	32.3	<LCL	n.d.	n.d.	n.d.
	Mean	59.4	n.c.	<LCL	n.d.	n.d.	n.d.
Constituent 6	1	41.2	27.6	<LCL	<LCL	<LCL	<LCL
	2	36.8	23.1	<LCL	<LCL	<LCL	<LCL
	Mean	39.0	25.4	<LCL	<LCL	<LCL	<LCL
Constituent 1	1	81.5	141.2	57.0	<LCL	111.6	n.d.
	2	113.1	105.3	92.5	<LCL	<LCL	<LCL
	Mean	97.3	123.3	74.8	<LCL	n.c.	<LCL
Constituent 7	1	45.3	34.7	<LCL	<LCL	<LCL	<LCL
	2	45.6	16.8	<LCL	<LCL	<LCL	<LCL
	Mean	45.5	25.7	<LCL	<LCL	<LCL	<LCL
Constituent 2	1	53.4	39.1	<LCL	<LCL	<LCL	<LCL
	2	56.3	24.4	n.d.	<LCL	<LCL	<LCL
	Mean	54.8	31.7	<LCL	<LCL	<LCL	<LCL
Constituent 3	1	58.8	34.5	<LCL	<LCL	11.9	<LCL
	2	60.7	22.8	n.d.	<LCL	<LCL	<LCL
	Mean	59.7	28.6	<LCL	<LCL	n.c.	<LCL
Constituent 4	1	69.8	50.0	12.8	10.5	16.0	<LCL
	2	74.0	33.7	16.0	<LCL	<LCL	<LCL
	Mean	71.9	41.8	14.4	n.c.	n.c.	<LCL
Constituent 5	1	65.3	54.1	20.9	20.8	19.4	n.d.
	2	71.0	43.9	19.4	22.4	<LCL	<LCL
	Mean	68.1	49.0	20.1	21.6	n.c.	<LCL

Repl. = Replicate; LCL = Lowest calibration level; n.d. = not detected; n.c.= not calculable 

Table 7. 100 µg/L test item

		% Applied (nominal)
Constituent	Repl.	Day
		0	2	5	8	12	14	20	33
Constituent 9	1	95.9	93.5	93.0	96.7	63.4	60.0	33.4	4.8*
	2	92.5	92.6	117.9	88.1	59.4	59.4	23.0	21.2
	Mean	94.2	93.0	105.4	92.4	61.4	59.7	28.2	13.0
Constituent 10	1	94.3	92.6	89.5	96.0	61.6	54.0	27.3	9.5*
	2	95.7	88.6	107.3	80.3	57.7	56.3	20.1	24.5
	Mean	95.0	90.6	98.4	88.2	59.7	55.2	23.7	17.0
Constituent 11	1	97.0	101.9	94.5	84.9	59.4	52.0	21.8	4.8*
	2	93.8	96.8	96.9	80.7	46.8	53.5	18.3	24.2
	Mean	95.4	99.4	95.7	82.8	53.1	52.8	20.1	14.5
Constituent 6	1	86.3	78.8	73.2	75.6	46.4	42.8	26.3	9.9
	2	83.1	81.6	81.7	65.2	40.9	44.8	17.9	28.5
	Mean	84.7	80.2	77.5	70.4	43.6	43.8	22.1	19.2
Constituent 1	1	99.3	93.7	97.5	103.3	81.6	95.5	79.1	81.6
	2	89.9	97.9	109.7	93.1	90.1	95.0	93.5	107.2
	Mean	94.6	95.8	103.6	98.2	85.8	95.2	86.3	94.4
Constituent 7	1	84.0	79.5	72.9	74.7	53.1	48.6	30.9	15.0
	2	83.7	83.3	77.9	65.6	44.7	50.7	22.5	32.5
	Mean	83.9	81.4	75.4	70.1	48.9	49.6	26.7	23.8
Constituent 2	1	89.0	92.9	84.8	85.4	54.6	48.9	28.3	8.7
	2	86.7	92.1	95.5	67.4	44.5	52.8	17.4	33.0
	Mean	87.8	92.5	90.1	76.4	49.6	50.9	22.9	20.9
Constituent 3	1	90.4	90.5	77.7	84.7	57.1	52.9	31.9	14.3
	2	85.6	89.0	91.1	67.6	49.5	53.8	19.2	31.1
	Mean	88.0	89.8	84.4	76.1	53.3	53.3	25.5	22.7
Constituent 4	1	89.4	90.3	81.7	83.8	66.9	63.1	47.6	27.4
	2	85.6	90.9	89.4	70.3	57.0	62.3	33.7	43.8
	Mean	87.5	90.6	85.5	77.1	61.9	62.7	40.7	35.6
Constituent 5	1	91.9	87.2	78.9	82.9	70.4	76.2	59.3	43.7
	2	85.9	85.6	91.0	70.3	61.0	66.5	45.2	60.9
	Mean	88.9	86.4	84.9	76.6	65.7	71.3	52.3	52.3

Table 7 (continued).100 µg/L test item

		% Applied (nominal)
Constituent	Repl.	Day
		43	51	57	62	75		90
Constituent 9	1	4.8*	4.8*	4.8*	4.8*	4.8*	4.8*	1.0*	1.0*
	2	4.8*	4.8*	4.8*	4.8*	9.1	4.8*	1.0*	1.0*
	Mean	4.8	4.8	4.8	4.8	6.9	4.8	1.0	1.0
Constituent 10	1	9.5	2.0	2.0	2.0	9.5*	9.5*	2.0*	2.0*
	2	9.5	2.0	2.0	2.0	12.0	9.5*	2.0*	2.0*
	Mean	9.5	2.0	2.0	2.0	10.8	9.5	2.0	2.0
Constituent 11	1	1.0*	1.0*	1.0*	1.0*	1.0*	1.0*	1.0*	1.0*
	2	1.0*	1.0*	1.0*	1.0*	25.8	1.0*	1.0*	1.0*
	Mean	1.0	1.0	1.0	1.0	13.4	1.0	1.0	1.0
Constituent 6	1	13.0	4.8	6.0	4.8	4.8	4.8	4.8*	4.8*
	2	10.1	4.8	4.8	4.8	12.8	7.2	4.8*	4.8*
	Mean	11.6	4.8	5.4	4.8	8.8	6.0	4.8	4.8
Constituent 1	1	85.8	30.6	103.9	53.7	6.0*	6.0*	6.0*	6.0*
	2	77.0	11.2	6.0	15.7	7.9	6.0*	6.0*	6.0*
	Mean	81.4	20.9	55.0	34.7	7.0	6.0	6.0	6.0
Constituent 7	1	15.1	8.2	6.0	4.8*	6.4	4.8*	4.8*	4.8*
	2	11.6	9.6	4.8*	4.8*	14.4	9.5	4.8*	4.8*
	Mean	13.3	8.9	5.4	4.8	10.4	7.1	4.8	4.8
Constituent 2	1	13.1	4.8*	5.9	4.8*	4.8*	4.8*	4.8*	4.8*
	2	6.3	4.8*	4.8*	4.8*	6.7	4.8*	4.8*	4.8*
	Mean	9.7	4.8	5.3	4.8	5.7	4.8	4.8	4.8
Constituent 3	1	18.0	4.8*	8.7	4.8*	4.8*	4.8*	4.8*	4.8*
	2	4.8*	4.8*	4.8*	4.8*	9.0	4.8*	4.8*	4.8*
	Mean	11.4	4.8	6.7	4.8	6.9	4.8	4.8	4.8
Constituent 4	1	38.6	7.2	22.1	6.8	4.8*	4.8*	4.8*	4.8*
	2	8.6	6.0	4.8*	4.8*	9.7	6.5	4.8*	4.8*
	Mean	23.6	6.6	13.4	5.8	7.2	5.6	4.8	4.8
Constituent 5	1	54.8	15.7	33.4	13.1	6.0	4.8*	4.8*	4.8*
	2	20.7	11.7	4.8*	4.8*	11.6	7.9	4.8*	4.8*
	Mean	37.8	13.7	19.1	8.9	8.8	6.3	4.8	4.8

Repl. = Replicate    *= value < lowest calibration level or not detected, respectively     

Table 8. Sterile Control - 20 µg/L

		% Applied (nominal)
Constituent	Repl.	Day
		0	2	5	8	12	14
Constituent 9	1	102.8	121.3	103.0	125.1	132.1	136.7
	2	84.4	109.7	129.5	106.9	118.5	111.9
	Mean	93.6	115.5	116.3	116.0	125.3	124.3
Constituent 10	1	108.3	119.3	113.4	137.7	143.9	139.6
	2	110.9	112.8	138.7	110.7	121.5	114.0
	Mean	109.6	116.0	126.0	124.2	132.7	126.8
Constituent 6	1	88.2	107.9	78.8	92.8	95.7	84.8
	2	90.6	98.7	103.0	87.2	88.7	76.5
	Mean	89.4	103.3	90.9	90.0	92.2	80.6
Constituent 1	1	127.5	124.3	120.1	137.3	131.8	124.1
	2	126.9	104.6	122.8	122.2	97.0	98.9
	Mean	127.2	114.4	121.4	129.7	114.4	111.5
Constituent 7	1	90.2	105.2	77.4	82.4	95.9	70.2
	2	91.4	95.9	101.7	77.2	88.2	68.6
	Mean	90.8	100.5	89.5	79.8	92.1	69.4
Constituent 2	1	103.9	118.8	90.7	107.4	119.0	116.6
	2	106.6	115.0	112.3	100.4	107.3	89.9
	Mean	105.2	116.9	101.5	103.9	113.2	103.2
Constituent 3	1	105.4	120.3	84.7	98.7	116.5	87.6
	2	100.6	111.2	106.2	98.8	95.3	77.8
	Mean	103.0	115.8	95.4	98.8	105.9	82.7
Constituent 4	1	96.8	117.9	78.0	88.3	95.6	78.2
	2	99.9	112.3	96.3	95.5	91.9	75.8
	Mean	98.3	115.1	87.2	91.9	93.8	77.0
Constituent 5	1	100.8	121.6	69.0	77.7	104.6	61.6
	2	105.2	116.7	89.3	86.5	78.0	53.2
	Mean	103.0	119.1	79.2	82.1	91.3	57.4

 

Table 8 (continued). Sterile Control - 20 µg/L

		% Applied (nominal)
Constituent	Repl.	Day
		20	33	43	51	57	62
Constituent 9	1	109.9	137.5	124.1	104.4	103.5	115.5
	2	111.8	132.0	122.0	103.6	110.6	96.3
	Mean	110.8	134.8	123.1	104.0	107.0	105.9
Constituent 10	1	102.9	141.1	107.0	99.4	105.3	108.7
	2	115.5	122.7	102.2	123.8	126.4	115.9
	Mean	109.2	131.9	104.6	111.6	115.8	112.3
Constituent 6	1	78.2	73.5	64.6	53.9	74.0	58.6
	2	115.9	70.3	47.2	69.9	75.6	65.8
	Mean	97.0	71.9	55.9	61.9	74.8	62.2
Constituent 1	1	85.8	146.9	91.1	164.7	138.0	129.5
	2	127.2	143.6	79.5	142.1	124.8	131.4
	Mean	106.5	145.3	85.3	153.4	131.4	130.4
Constituent 7	1	78.0	74.6	55.0	53.9	64.2	49.3
	2	117.2	51.7	31.6	57.4	66.3	61.6
	Mean	97.6	63.2	43.3	55.6	65.3	55.4
Constituent 2	1	99.8	107.8	81.9	100.1	111.2	107.3
	2	146.0	87.7	61.5	100.8	108.4	104.4
	Mean	122.9	97.7	71.7	100.4	109.8	105.8
Constituent 3	1	92.8	96.5	60.5	85.3	91.0	88.2
	2	126.1	71.8	32.1	81.4	96.0	87.9
	Mean	109.4	84.2	46.3	83.4	93.5	88.1
Constituent 4	1	80.1	85.4	38.3	73.1	68.9	71.9
	2	130.7	55.9	24.4	67.5	78.4	66.9
	Mean	105.4	70.6	31.3	70.3	73.6	69.4
Constituent 5	1	71.1	71.7	28.3	63.4	52.1	55.6
	2	110.8	37.4	20.0	69.3	56.9	67.2
	Mean	91.0	54.6	24.2	66.4	54.5	61.4

Repl. = Replicate 

Table 9. Sterile Control - 100 µg/L

		% Applied (nominal)
Constituent	Repl.	Day
		0	2	5	8	12	14	20	33
Constituent 9	1	94.8	97.8	101.4	103.0	94.6	105.2	106.9	106.3
	2	95.0	96.7	98.9	99.7	96.1	108.9	104.1	113.7
	Mean	94.9	97.2	100.1	101.4	95.4	107.0	105.5	110.0
Constituent 10	1	94.1	103.8	94.0	95.8	92.0	101.4	100.3	105.5
	2	98.6	98.9	97.4	101.2	99.3	109.0	101.7	115.0
	Mean	96.4	101.4	95.7	98.5	95.6	105.2	101.0	110.2
Constituent 11	1	92.2	101.5	92.9	95.9	87.7	100.6	99.3	115.2
	2	104.8	94.5	106.0	93.6	98.3	93.2	105.8	105.6
	Mean	98.5	98.0	99.4	94.8	93.0	96.9	102.6	110.4
Constituent 6	1	81.8	84.0	81.6	80.6	79.7	78.7	89.8	70.6
	2	84.5	80.6	84.2	91.4	79.9	75.8	82.8	70.7
	Mean	83.2	82.3	82.9	86.0	79.8	77.3	86.3	70.7
Constituent 1	1	105.1	101.2	103.9	107.7	96.4	101.3	106.8	114.6
	2	99.8	97.3	97.4	104.2	105.2	101.0	110.8	106.0
	Mean	102.5	99.2	100.6	106.0	100.8	101.1	108.8	110.3
Constituent 7	1	79.7	83.3	78.0	74.2	74.3	71.1	88.3	66.2
	2	80.8	77.8	85.0	90.9	77.6	66.1	76.0	63.0
	Mean	80.3	80.5	81.5	82.6	76.0	68.6	82.2	64.6
Constituent 2	1	94.9	104.4	93.6	95.9	97.6	94.9	111.3	92.1
	2	92.4	101.3	100.2	103.8	94.2	93.3	103.2	86.1
	Mean	93.7	102.8	96.9	99.9	95.9	94.1	107.2	89.1
Constituent 3	1	88.9	103.5	88.6	85.4	89.6	85.7	102.7	82.3
	2	90.5	91.3	91.8	99.3	91.5	79.4	96.4	76.9
	Mean	89.7	97.4	90.2	92.3	90.6	82.5	99.6	79.6
Constituent 4	1	89.3	99.3	85.3	77.7	85.5	78.3	94.7	76.9
	2	90.8	91.5	92.5	94.9	81.8	69.0	85.3	72.5
	Mean	90.0	95.4	88.9	86.3	83.7	73.6	90.0	74.7
Constituent 5	1	86.0	94.1	89.2	77.4	80.6	67.9	90.3	74.1
	2	86.7	87.0	99.8	97.8	80.9	58.4	81.4	66.1
	Mean	86.3	90.6	94.5	87.6	80.8	63.1	85.9	70.1

Table 9 (continued). Sterile Control - 100 µg/L

Constituent	Repl.	% Applied (nominal)
		Day
		43	51	57	62	75	90
Constituent 9	1	98.8	97.5	97.6	97.9	91.2	88.4
	2	103.0	99.9	100.6	102.9	114.5	93.6
	Mean	100.9	98.7	99.1	100.4	102.9	91.0
Constituent 10	1	101.0	95.0	102.9	103.6	92.7	77.8
	2	98.5	98.7	102.4	105.7	114.6	95.1
	Mean	99.7	96.8	102.6	104.6	103.7	86.5
Constituent 11	1	98.2	97.9	91.5	102.5	68.4	77.5
	2	115.0	98.9	110.8	100.0	120.7	102.8
	Mean	106.6	98.4	101.2	101.3	94.6	90.2
Constituent 6	1	72.4	46.8	46.0	36.5	29.2	20.4
	2	80.6	59.8	59.6	59.6	58.2	50.2
	Mean	76.5	53.3	52.8	48.1	43.7	35.3
Constituent 1	1	98.8	101.6	90.7	68.4	63.6	50.3
	2	117.3	104.7	75.7	91.9	89.4	87.2
	Mean	108.1	103.1	83.2	80.1	76.5	68.8
Constituent 7	1	67.9	40.1	39.0	30.4	20.8	15.9
	2	73.2	58.0	54.6	54.9	53.1	44.8
	Mean	70.5	49.0	46.8	42.6	36.9	30.3
Constituent 2	1	97.5	69.8	75.8	68.1	48.0	34.3
	2	104.6	79.8	82.6	89.4	70.7	66.2
	Mean	101.0	74.8	79.2	78.7	59.3	50.3
Constituent 3	1	90.9	61.0	62.9	54.7	39.6	31.1
	2	95.8	71.1	70.1	70.8	57.6	56.4
	Mean	93.3	66.0	66.5	62.7	48.6	43.8
Constituent 4	1	79.1	58.0	55.5	49.8	39.8	29.9
	2	82.9	65.2	59.6	60.5	54.5	49.6
	Mean	81.0	61.6	57.5	55.2	47.1	39.7
Constituent 5	1	74.4	52.9	52.1	47.2	40.9	31.0
	2	70.9	57.6	54.6	53.6	56.4	45.3
	Mean	72.6	55.2	53.4	50.4	48.6	38.1

Repl. = Replicate

Table 10. Data of Kinetic Fit and DTx Values of the test item - 20 μg/L

Endpoint / Statistic	Single First Order (SFO)
Constituent	Constituent 9	Constituent 10	Constituent 6	Constituent 7
C0	122.6 ± 6.179	121.6 ± 5.657	88.34 ± 2.389	86.68 ± 3.881
Initial value for fitting	100	100	100	100
K1 (1/d)	0.03545 ± 0.005252	0.03272 ± 0.005029	0.03565 ± 0.002825	0.03186 ± 0.004459
Initial value for fitting	0.1	0.1	0.1	0.1
r2	0.842	0.8208	0.9435	0.8393
c2 error (parent)	9.47	7.82	4.64	5.30
	DTx Values in days
DT50	19.6	21.2	19.4	21.8
Endpoint / Statistic	Single First Order (SFO)
Constituent	Constituent 2	Constituent 3	Constituent 4	Constituent 5
C0	109.0 ± 3.504	100.8 ± 2.810	104.3 ± 4.366	105.5 ± 3.51
Initial value for fitting	100	100	100	100
K1 (1/d)	0.03538 ± 0.003349	0.03083 ± 0.002613	0.02885 ± 0.003258	0.02753 ± 0.002415
Initial value for fitting	0.1	0.1	0.1	0.1
r2	0.9205	0.9462	0.8995	0.9281
c2 error (parent)	4.78	5.64	12.1	7.07
	DTx Values in days
DT50	19.6	22.5	24.0	25.2

 

Table 11. Data of Kinetic Fit and DTx Values of the test item - 100 µg/L

Endpoint / Statistic	Single First Order (SFO)
Constituent	Constituent 9	Constituent 10	Constituent 11	Constituent 6	Constituent 1
C0	110.7 ± 4.971	107.1 ± 4.513	110.4 ± 4.752	89.83 ± 3.081	111.4 ± 8.292
Initial value for fitting	100	100	100	100	100
K1 (1/d)	0.004996 ± 0.004922	0.005047 ± 0.004662	0.05832 ± 0.005457	0.05007 ± 0.003766	0.01857 ± 0.003106
Initial value for fitting	0.1	0.1	0.1	0.1	0.1
r2	0.9343	0.9402	0.9432	0.9588	0.7066
c2 error (parent)	19.6	17.9	17.8	12.7	20.6
	DTx Values in days
DT50	13.9	13.7	11.9	13.8	37.3
DT90	46.1	45.6	39.5	46	124
Endpoint / Statistic	Single First Order (SFO)
Constituent	Constituent 7	Constituent 2	Constituent 3	Constituent 4	Constituent 5
C0	88.18 ± 2.564	99.75 ± 3.903	97.27 ± 3.336	65.80 ± 3.173	94.28 ± 3.747
Initial value for fitting	100	100	100	100	100
K1 (1/d)	0.04287 ± 0.002735	0.05061 ± 0.004342	0.04673 ± 0.003519	0.03524 ± 0.002539	0.02748 ± 0.002356
Initial value for fitting	0.1	0.1	0.1	0.1	0.1
r2	0.9668	0.9471	0.9569	0.9522	0.9212
c2 error (parent)	9.96	14.8	12.2	9.18	10.9
	DTx Values in days
DT50	16.2	13.7	14.8	19.7	25.2
DT90	53.7	45.5	49.3	65.3	83.8

 

Table12. Identified metabolites

Metabolite (M)	Molecular Formula	Retention Time [min]	Accurate Mass Adduct [M+H]+ m/z	Collision Energy [V]
4	C26H43NO5	6.58	450.3214	20
5	C26H45NO5	6.53	452.3371	20
9	C29H51NO6	6.66	510.3789	20
10	C31H54N2O7	5.47	567.4004	20
11	C31H56N2O7	5.44	569.4160	20
20	C34H58N2O8	5.53	623.4266	20, 30
13	C34H60N2O8	5.51	625.4422	20
14	C34H62N2O8	5.48	627.4579	20, 30
17	C37H64N2O9	5.59	681.4685	30
18	C37H66N2O9	5.54	683.4841	30
19	C37H68N2O9	5.53	685.4998	30

Table 13. Link between Metabolites and corresponding Constituents of the Test Item

           DBE = Double Bond Equivalents

	Constituent	Metabolite
No.	6	M4	M5
DBE	4	6	5
Mass Fragment Ions [amu]	291.2319	289.2162 261.2213	291.2319
Fragment Ions	[C19H31O2]+	[C19H29O2]+ [C18H29O]+	[C19H31O2]+
No.	7	M9	--
DBE	4	5	--
Mass Fragment Ions [amu]	291.2319 275.2369	289.2162	--
Fragment Ions	[C19H31O2]+ [C19H31O]+	[C19H29O2]+	--
	Constituent	Metabolite
No.	2	M10		M11		--
DBE	4	6		5		--
Mass Fragment Ions [amu]	466.3527 408.3108 303.2319	464.3371 303.2319		464.3371 408.3108 303.2319		--
Fragment Ions	[C27H48NO5]+ [C24H42NO5]+ [C20H31O2]+	[C27H46NO5]+ [C20H31O2]+		[C27H46NO5]+ [C24H42NO4]+ [C20H31O2]+		--
No.	3	M20		M13		M14
DBE	4	7		6		5
Mass Fragment Ions [amu]	524.3946 466.3527 303.2319	464.3371 303.2319		464.3371 303.2319		466.3527 464.3371 303.2319
Molecular Formula Fragment Ions	[C30H54NO6]+ [C27H48NO5]+ [C20H31O2]+	[C27H46NO5]+ [C20H31O2]+		[C27H46NO5]+ [C20H31O2]+		C27H48NO5]+ [C27H46NO5+ [C20H31O2]+
No.	4	M17		M18		M19
DBE	4	7		6		5
Mass Fragment Ions [amu]	524.3946 466.3527 303.2319	303.2319		522.3789 464.3371 303.2319		522.3789 303.2319
Fragment Ions	[C30H54NO6]+ [C27H48NO5]+ [C20H31O2]+	[C20H31O2]+		[C30H52NO6]+ [C27H46NO5]+ [C20H31O2]+-		[C30H52NO6]+ [C20H31O2]+

Table 14 Data of Kinetic Fit and DT_xValues of Metabolites100 µg/L

Endpoint / Statistic	100 µg/L
Model	Single First Order (SFO)
Metabolite	M4	M5	M9	M10	M11	M13
C0	111.2 ± 10.48	117.5 ± 17.20	115.0 ± 21.86	103.5 ± 14.87	96.13 ± 18.43	97.08 ± 15.49
Initial value for fitting	100	100	100	100	100	100
K1 (1/d)	0.05278 ± 0.008055	0.03289 ± 0.007743	0.03432 ± 0.01163	0.05671 ± 0.01504	0.08724 ± 0.03072	0.09153 ± 0.02702
Initial value for fitting	0.1	0.1	0.1	0.1	0.1	0.1
r2	0.8539	0.6742	0.5601	0.7259	0.6278	0.7129
c2 error (parent)	23.6	29.5	35.7	19.6	27.7	21.1
	DTx Values in days
DT50	13.1	21.1	20.2	12.2	7.95	7.57
DT90	43.6	70	67.1	40.6	26.4	25.2
Endpoint / Statistic	100 µg/L
Model	Single First Order (SFO)
Metabolite	M14	M17	M18	M19	M20
C0	111.1 ± 19.09	99.01 ± 11.64	98.12 ± 12.67	110.5 ± 18.34	110.2 ± 18.39
Initial value for fitting	100	100	100	100	100
K1 (1/d)	0.05111 ± 0.01475	0.08286 ± 0.01779	0.08867 ± 0.02108	0.04813 ± 0.01347	0.05065 ± 0.01596
Initial value for fitting	0.1	0.1	0.1	0.1	0.1
r2	0.6403	0.8188	0.7913	0.6472	0.6472
c2 error (parent)	28.8	12.8	14.8	23.1	24.4
DT50	13.6	8.37	7.82	14.4	13.7
DT90	45.1	27.8	26.0	47.8	45.5

Validity criteria:

The reference item aniline was degraded (> 50%) within 6 days.

Observed value:

The reference item aniline was degraded 60% within 6 days.

Validity criteria fulfilled:

yes

Conclusions:

In a biodegradation simulation study that was performed in accordance with OECD TG 309, the DT50 values of the test item at 20 µg/L applied concentration in surface water were in a range of 19.4 to 25.2 days. The DT50 values of the test item at 100 µg/L applied concentration in surface water were in a range of 11.9 to 37.3 days. Metabolites were formed by oxidation of the primary or secondary alcohol moieties of the constituents. Eleven metabolites (M) were identified in the test. The DT50 values of the metabolites of the test item at 100 µg/L applied concentration in surface water were in a range of 7.57 to 14.4 days.

Executive summary:

The aerobic mineralization of the test item in surface water was determined in a simulation biodegradation test over a test period of 90 days according to OECD TG 309 and in compliance with GLP criteria. The study was carried out at 12 ± 2 °C, pH 7.50 – 7.85, 9.29 – 9.60 mg O2/L oxygen concentration in darkness. Duplicate per sample per concentration for test item and sterile controls were included in the test. In addition, one blank control, one sterile blank control, two reference controls and two solvent controls (methanol) were included in the experimental set-up as well. Sampling for determination of the transformation was carried out directly after application and at 13 additional sampling points (day 2, 5, 8, 12, 14, 20, 33, 43, 51, 57, 62, 75 and day 90). The amount of test item constituents and degradation products in the test solution and sterile test item solution was determined by LC-HRMS. Residual activity in the test solution and dissolved 14CO2 (after acidification of test solution) was determined by LSC.

The test item is a UVCB, a reaction product of p-nonylphenol, formaldehyde and diethanolamine, propoxylated. Thus, it cannot be radiolabelled, in turn, analysis of Non-extractable residues (NER) was not possible. Eleven constituents (target compounds) were known. The results showed that transformation of most constituents started in within 5 – 12 days after application. About 50 % transformation was reached within 33 - 43 days, after 62 days the detected concentration was < LOQ or even not detectable. For constituent 1 transformation started only on day 43 and the transformation reached 50% after 57 days and 65 % on day 62. At day 75 the concentration was < LOQ. Metabolites were formed by oxidation of the primary or secondary alcohol moieties of the constituents. Eleven metabolites (M) were identified in the test. M4 and M5 were identified in Constituent 6 transformation products. M9 was identified in Constituent 7 transformation products. M10 and M11 were identified in Constituent 2 transformation products. M13, M14 and M20 were identified in Constituent 3 transformation products. M17, M18 and M19 were identified in Constituent 4 transformation products.

Based on the findings, the DT50 values of the test item at 20 µg/L applied concentration in surface water was in a range of 19.4 to 25.2 days. The DT50 values of the test item at 100 µg/L applied concentration in surface water were in a range of 11.9 to 37.3 days. The DT50 values of the metabolites of the test item at 100 µg/L applied concentration in surface water was in a range of 7.57 to 14.4 days.

Description of key information

The DT50 values of the test item at 20 µg/L applied concentration in surface freshwater were in a range of 19.4 to 25.2 days. The DT50 values of the test item at 100 µg/L applied concentration in surface freshwater were in a range of 11.9 to 37.3 days. The DT50 values of the metabolites of the test item at 100 µg/L applied concentration in surface freshwater were in a range of 7.57 to 14.4 days.

Key value for chemical safety assessment

Half-life in freshwater:

37.3 d

at the temperature of:

12 °C

Additional information

The aerobic mineralization of the test item in surface water was determined in a simulation biodegradation test over a test period of 90 days according to OECD TG 309 and in compliance with GLP criteria. The study was carried out at 12 ± 2 °C, pH 7.50 – 7.85, 9.29 – 9.60 mg O2/L oxygen concentration in darkness. Duplicate per sample per concentration for test item and sterile controls were included in the test. In addition, one blank control, one sterile blank control, two reference controls and two solvent controls (methanol) were included in the experimental set-up as well. Sampling for determination of the transformation was carried out directly after application and at 13 additional sampling points (day 2, 5, 8, 12, 14, 20, 33, 43, 51, 57, 62, 75 and day 90). The amount of test item constituents and degradation products in the test solution and sterile test item solution was determined by LC-HRMS. Residual activity in the test solution and dissolved 14CO2 (after acidification of test solution) was determined by LSC.

The test item is a UVCB, a reaction product of p-nonylphenol, formaldehyde and diethanolamine, propoxylated. Thus, it cannot be radiolabelled, in turn, analysis of Non-extractable residues (NER) was not possible. Eleven constituents (target compounds) were known. The results showed that transformation of most constituents started in within 5 – 12 days after application. About 50 % transformation was reached within 33 - 43 days, after 62 days the detected concentration was < LOQ or even not detectable. For constituent 1 transformation started only on day 43 and the transformation reached 50% after 57 days and 65 % on day 62. At day 75 the concentration was < LOQ. Metabolites were formed by oxidation of the primary or secondary alcohol moieties of the constituents. Eleven metabolites (M) were identified in the test. M4 and M5 were identified in Constituent 6 transformation products. M9 was identified in Constituent 7 transformation products. M10 and M11 were identified in Constituent 2 transformation products. M13, M14 and M20 were identified in Constituent 3 transformation products. M17, M18 and M19 were identified in Constituent 4 transformation products.

Based on the findings, the DT50 values of the test item at 20 µg/L applied concentration in surface water was in a range of 19.4 to 25.2 days. The DT50 values of the test item at 100 µg/L applied concentration in surface water were in a range of 11.9 to 37.3 days. The DT50 values of the metabolites of the test item at 100 µg/L applied concentration in surface water was in a range of 7.57 to 14.4 days.