Ethyl [2-(4-phenoxyphenoxy)ethyl]carbamate

Biodegradation in soil

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in soil: simulation testing

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

28 Feb 1994 to 31 Mar 1995

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: EPA 162-1 and 162-2

Deviations:

yes

Remarks:

Mass balance = 82 - 89% AR (kinetic study) from 2 months onwards, but the results were considered sufficiently reliable.

GLP compliance:

yes (incl. QA statement)

Test type:

laboratory

Radiolabelling:

yes

Oxygen conditions:

aerobic/anaerobic

Soil classification:

USDA (US Department of Agriculture)

Year:

1993

Soil no.:

#1

Soil type:

sandy loam

% Clay:

10

% Silt:

16

% Sand:

74

% Org. C:

1.3

pH:

7.7

CEC:

10.9 meq/100 g soil d.w.

Bulk density (g/cm³):

1.24

% Moisture content:

10.4

Soil No.:

#1

Duration:

>= 360 - <= 365 d

Soil No.:

#1

Initial conc.:

0.12 mg/kg soil d.w.

Based on:

test mat.

Remarks:

Aerobic; kinetic study

Soil No.:

#1

Initial conc.:

9.2 mg/kg soil d.w.

Based on:

test mat.

Remarks:

Aerobic; degradate study

Soil No.:

#1

Initial conc.:

0.13 mg/kg soil d.w.

Based on:

other:

Remarks:

Anaerobic; kinetic study

Soil No.:

#1

Initial conc.:

9.9 mg/kg soil d.w.

Based on:

other: anaerobic; degradate study

Parameter followed for biodegradation estimation:

CO2 evolution

radiochem. meas.

Soil No.:

#1

Temp.:

25°C

Humidity:

75% of moisture capacity at 1/3 bar

Soil No.:

#1

% Total extractable:

76

% Non extractable:

25

% Recovery:

100

Remarks on result:

other: 6 hour; 0.12 mg/kg (aerobic; hourly kenetic)

Soil No.:

#1

% Total extractable:

4.8

% Non extractable:

41

% CO2:

38

% Recovery:

84

Remarks on result:

other: day 365; 0.12 mg/kg (aerobic; kinetic)

Soil No.:

#1

% CO2:

32

% Recovery:

89

Remarks on result:

other: day 365; 9.2 mg/kg (aerobic; degradate)

Soil No.:

#1

% Total extractable:

14

% Non extractable:

60

% CO2:

32

% Recovery:

100

Remarks on result:

other: day 360; 0.13 mg/kg (anaerobic; kinetic)

Soil No.:

#1

% CO2:

15

% Recovery:

80

Remarks on result:

other: 12 mo;9.9 mg/kg (anaerobic; degradate)

Parent/product:

parent

Soil No.:

#1

% Degr.:

39

Parameter:

radiochem. meas.

Sampling time:

6 h

Remarks on result:

other: 0.12 mg/kg (aerobic; hours kinetic study)

Parent/product:

parent

Soil No.:

#1

% Degr.:

80

Parameter:

radiochem. meas.

Sampling time:

1 d

Remarks on result:

other: 0.12 mg/kg (aerobic; kinetic study)

Parent/product:

parent

Soil No.:

#1

% Degr.:

72

Parameter:

radiochem. meas.

Sampling time:

14 d

Remarks on result:

other: 0.13 mg/kg (anearobic; kinetic study)

Key result

Soil No.:

#1

DT50:

81.5 d

Type:

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

Temp.:

25 °C

Remarks on result:

other: aerobic; calculated outside of the original study

Soil No.:

#1

DT50:

6.7 h

Type:

other: primary

Temp.:

25 °C

Remarks on result:

other: aerobic; reported in the original study (Thede 1995)

Soil No.:

#1

DT50:

8.2 mo

Type:

other: secondary

Temp.:

25 °C

Remarks on result:

other: aerobic; reported in the original study (Thede 1995)

Soil No.:

#1

DT50:

16 d

Type:

other: primary

Temp.:

25 °C

Remarks on result:

other: anearobic; reported in the original study (Thede 1995)

Soil No.:

#1

DT50:

8.5 mo

Type:

other: Secondary

Temp.:

25 °C

Remarks on result:

other: anearobic; reported in the original study (Thede 1995)

Transformation products:

not specified

Remarks:

M2, M4, M7/M10, M8 and M9

Evaporation of parent compound:

not specified

Volatile metabolites:

yes

Remarks:

CO2

Residues:

yes

Aerobic:

- Microbial activity: Microbial activity of the test soil was confirmed at the start, during and the end of aerobic incubation by determining the microbial biomass (plate count method). Soils were found to be viable throughout the incubation period.

- Recovery of radioactivity: Total recovery of radioactivity ranged from 82 to 99% AR (kinetic study) and from 95 to 102% AR (hourly kinetic study). The amount of soil extractable radioactivity decreased from 71% AR on day 0 to 4.8% AR after 1 year and from 97 to 76% AR over the 6 h period in the hourly kinetic study. Unextractable residues increased from 28% AR on day 0 to 65% AR on day 7 and declined again thereafter to 41% AR after 1 year; in the hourly kinetic study unextractable residues increased from 5.8 to 25% AR over the 6h period. CO2 was evolved from the treated soil to a maximum of 38% AR after 1 year. The distribution between soil radioactivity and CO2 (degradate study) confirms the CO2 evolution: 32% AR after one year.

- Degradation: The test substance degraded to 3.9% AR after 1 year with a fast decline in the first two days (to 12% AR). The hourly kinetic study showed a decline to 61% AR in the 6h period. At least 10regions of radioactivity (single fraction ≤ 3.6% AR) including 5 identified metabolites, were observed (M2 (max 3.3% AR), M4 (max 0.5% AR), M7/M10 (unresolved, max 0.15% AR), M8 (max 0.22% AR) and M9 (max 0.1% AR)).  

Anaerobic:

- Microbial activity: Microbial activity of the test soil was confirmed at the start, during and the end of aerobic incubation by determining the microbial biomass (plate count method). Soils and water were found to be viable throughout the incubation period.

- recovery radioactivity: Total recovery of radioactivity ranged from 94 to 107% AR. The amount of soil extractable radioactivity decreased from 69% AR on day 0 to 14% AR after 1 year. Unextractable residues increased from 25% AR on day 0 to 60% AR after 9 months and remained stable thereafter. Radioactivity in the water layer was≤4.2% AR. CO2 was evolved to a maximum of 32% AR after 1 year.

The distribution between soil radioactivity and CO2 (degradate study) showed a lower CO2 evolution: 15% AR after one year.

- Degradation: The test substance degraded to 8.1% AR after 1 year with a fast decline in the first two weeks (to 28% AR). At least 10 regions of radioactivity (≤4.3% AR) including 5 identified metabolites, were observed (M2 (max 3.3% AR), M4 (max 0.4% AR), M7/M10 (unresolved, max 0.16% AR), M8 (max 0.5% AR) and M9 (max 0.6% AR)).

Table 2. Distribution of radioactivity after incubation at 25°C of sandy loam soil treated with [U-14C-phenoxyphenoxy]-labelled test substance at 0.12 mg/kg (kinetic study) (% AR, mean of duplicate samples)

time (d)	Extractables (cold extract)		unextractables			CO2	mass balance
	Total 1	Test substance	total	NaOH Extract 2	Soil combustion
0.125*	71	57	28	13	16	n.a.	99
0.25	50	36	43	17	26	2.4	95
0.5	38	28	51	27	24	4.5	93
1	29	20	54	30	24	7.7	91
1.25	24	17	62	30	32	8.6	94
2	23	12	62	32	31	10	96
3	19	14	64	34	30	12	95
7	14	10	65	39	25	15	93
14	12	8.8	62	23	39	14	89
21	11	8.3	63	26	37	19	93
30	9.0	7.1	63	22	40	21	92
59	7.1	5.6	57	21	36	25	89
88	6.2	5.0	53	17	36	29	89
182	4.8	3.8	47	17	30	36	88
273	3.8	3.1	42	15	27	36	82
365	4.8	3.9	41	23	18	38	84

1 including at least 10 regions of radioactivity (no single fraction >3.6% AR) with 5 identified metabolites

2 radioactivity considered to be associated with humic and fulvic acid; radioactivity could not be identified

* based on the time elapsed between treatment and sampling, 3h is considered more appropriate than 0 h

n.a.: not analysed

 

Table 3. Distribution of radioactivity after incubation at 25°C of sandy loam soil treated with [U-14C-phenoxyphenoxy]-labelled test substance at 0.12 mg/kg (hourly kinetic study) (% AR, mean of duplicate samples)

time (h)	extractables (cold extract)		unextractables			mass balance3
	Total 1	The test substance	total	NaOH Extract 2	soil combustion
0	97	95	5.8	1.2	4.6	102
1	92	80	8.6	2.4	6.2	100
2	88	86	13	4.7	8.1	101
3	88	78	14	6.3	7.4	102
4	79	66	17	5.4	11	95
5	77	68	19	5.7	13	96
6	76	61	25	8.2	17	101

1. including at least 10 regions of radioactivity (no single fraction > 1.9 % AR) with 5 identified metabolites

2 radioactivity considered to be associated with humic and fulvic acid; radioactivity could not be identified

3 No trapping of volatiles

 

Table 4. Distribution of radioactivity after incubation at 25°C of sandy loam soil treated with [U-14C-phenoxyphenoxy]-labelled test substance at 9.2 mg/kg (degradate study) (% AR, mean of duplicate samples).

Time (m)	0	0.5	1	3	6	9	12
Soil	100	83	79	69	65	58	57
CO2	n.a.	12	18	25	27	35	32
Mass balance	100	95	97	94	92	93	89

n.a.: not analysed

 

Table 5. Distribution of radioactivity after anaerobic incubation at 25°C of sandy loam soil treated with [U-14C-phenoxyphenoxy]-labelled test substance at 0.13 mg/kg (kinetic study) (% AR, mean of duplicate samples)

time (d)	extractables (cold extract)		unextractables			CO2	water layer	mass balance
	Total (A)	Test substance	total	NaOH Extract (B)	Soil combustion
0.125	69	60	25	6.7	19	n.p.	n.a.	94
0.25	56	44	36	8.3	28	12	3.6	107
14	41	28	44	19	25	17	4.2	106
30	31	21	50	15	35	21	1.9	104
90	23	15	52	16	36	25	1.4	101
180	18	12	60	17	42	20	2.3	100
360	14	8.1	60	24	36	32	1.9	107

(A) including at least 10 regions of radioactivity (individual fractions £4.3% AR) with 5 identified metabolites

(B) radioactivity considered to be associated with humic and fulvic acid; radioactivity could not be identified

n.p.: not performed

n.a.: not applicable

 

Table 6. Distribution of radioactivity after anaerobic incubation at 25°C of sandy loam soil treated with [U-14C-phenoxyphenoxy]-labelled test substance at 9.9 mg/kg (degradate study) (% AR, mean of duplicate samples)

Time (months)	1	6	9	12 (A)
soil	82	77	79	64
Water layer	3.9	2.2	2.0	1.3
CO2	7.7	14	13	15
Mass balance	94	93	94	80

(A) results of one sample

 

Conclusions:

In a biodegradation study in Buckeystown Soil (sandy loam), performed in accordance with EPA guideline 162-1 and 162-2, the degradation of the test substance is characterised by a very rapid initial decline, followed by a slow phase. Degradation of the test substance for the aerobic and anaerobic kinetic phases followed biphasic kinetics. The primary half-life for the aerobic kinetic incubation was determined to be 6. 7 hours and the secondary half-life value was calculated to be 8.2 months. The anaerobic kinetic incubation primary half-life was calculated to be 16 days and the secondary half-life value was calculated to be 8.5 months. The DT50 value in aerobic incubation was re-calculated in SFO model outside of the original study, and it was 81.5 days.

Executive summary:

The biodegradation of the[U-14C-phenoxyphenoxy]-labelled test substance has been investigated in aerobic and anaerobic studies. The studies were in accordance with EPA guideline 162-1 and 162-2, and in compliance with GLP criteria. The Buckeystownsoil (sandy loan, pH 7.7) was treated with the 14C-labelled test substance at a rate of 0.12 (aerobic; kinetic study), 9.2 (aerobic; degradate study), 0.13 (anaerobic; kinetic study) or 9.9 (anaerobic; degradate study) mg/kg dry soil. The treated soil were adjusted to 75% of moisture capacity at 1/3 bar and incubated at 25 °C in the dark for up to 1 year. The studied were carried out in accordance with EPA guideline 162-1 and 162-2, and in compliance with GLP criteria. The soil moisture content was maintained at the initial level. Aerobic conditions were maintained by passing humidified air through the incubation flasks for ~30 min at each sampling interval or every two weeks (whichever was shorter). Outgoing air was passed through two 10% KOH traps for CO2 trapping. No other traps for volatiles were used. The aerobic kinetic study was extended with an additional set of 20 g dry weight soil samples, fortified individually at a rate of 0.12 mg/kg (using 10 μL acetonitrile) and incubated under the same conditions for up to 6 hours, without trapping of volatiles (hourly kinetic study).

For aerobic studies, duplicate soil samples were analysed after 0, 1, 2, 3, 4, 5 and 6 hours (hourly kinetic study); 0, 3, 6, 12, 24, 30 hours, 2, 3, 7, 14, 21 days, 1, 2, 3, 6, 9 and 12 months (kinetic study); 6 hours, 14 days, 1, 3, 6, 9 and 12 months (degradate study) post treatment. Total radioactivity in soil was determined by combustion/LSC and in trapping solutions by LSC (except for hourly kinetic samples). For anaerobic studies, duplicate soil samples were analysed after 3, 6 hours, 14 days, 1, 3, 6 and 12 months (kinetic study); 1, 6, 9 and 12 (one sample) months (degradate study) post treatment. Total radioactivity in soil was determined by combustion/LSC; in the water layer and the trapping solutions by LSC. The test substancewas quantified by 1D-TLC and HPLC (both methods yielded similar results) and identity was confirmed by co-chromatography with reference standards.

For aerobic studies, the total recovery of radioactivity ranged from 82 to 99% AR (kinetic study), from 95 to 102% AR (hourly kinetic study) and from 89 to 100% AR (degradated study). The test substance degraded to 3.9% AR after 1 year with a fast decline in the first two days (to 12% AR). The hourly kinetic study showed a decline to 61% AR in the 6h period. At least 10 regions of radioactivity (single fraction ≤ 3.6% AR) including 5 identified metabolites (i.e. M2, M4, M7/M10, M8 and M9). For anaerobic studies, the total recovery of radioactivity ranged from 94 to 107% AR.The test substancedegraded to 8.1% AR after 1 year with a fast decline in the first two weeks (to 28% AR). At least 10 regions of radioactivity ( ≤ 4.3% AR) including 5 identified metabolites (same as in aerobic studies).

The degradation of the test substance is characterised by a very rapid initial decline, followed by a slow phase. Degradation of the test substance for the aerobic and aerobic/anaerobic kinetic phases followed biphasic kinetics. The primary half-life for the aerobic kinetic incubation was determined to be 6. 7 hours and the secondary half-life value was calculated to be 8.2 months. The SFO kenetic model recalculated DT50 is 81.5 days. The anaerobic kinetic incubation primary half-life was calculated to be 16 days and the secondary half-life value was calculated to be 8.5 months. The DT50 value in aerobic incubation was re-calculated in SFO model outside of the original study, and it was 81.5 days.

Reference 2

Endpoint:

biodegradation in soil: simulation testing

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

01 Feb 1993 to 04 Mar 1994

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: EPA 162-1 and EPA 162-2

Deviations:

not specified

GLP compliance:

yes (incl. QA statement)

Test type:

laboratory

Radiolabelling:

yes

Oxygen conditions:

aerobic/anaerobic

Soil classification:

USDA (US Department of Agriculture)

Year:

1993

Soil no.:

#1

Soil type:

sandy loam

% Clay:

17

% Silt:

12

% Sand:

71

% Org. C:

1.5

pH:

7.8

CEC:

10.6 meq/100 g soil d.w.

Bulk density (g/cm³):

1.16

% Moisture content:

17.3

Soil No.:

#1

Duration:

12 mo

Soil No.:

#1

Initial conc.:

>= 10.23 - <= 10.55 mg/kg soil d.w.

Based on:

test mat.

Parameter followed for biodegradation estimation:

CO2 evolution

radiochem. meas.

Soil No.:

#1

Temp.:

25 ± 1°C

Microbial biomass:

75% of field moisture capacity (1/3 bar)

Soil No.:

#1

% Total extractable:

1.8

% Non extractable:

58

% CO2:

31

% Recovery:

91

Remarks on result:

other: 212d; 10.23 mg/kg; aerobic

Soil No.:

#1

% Total extractable:

21

% Non extractable:

44

% CO2:

20

% Recovery:

85

Remarks on result:

other: 59d; 10.55 mg/kg; aerobic

Soil No.:

#1

% Total extractable:

16

% Non extractable:

69

% CO2:

20

% Other volatiles:

0

% Recovery:

100

Remarks on result:

other: 0 d; anaerobic

Soil No.:

#1

% Non extractable:

58

% CO2:

26

% Other volatiles:

0

% Recovery:

85

Remarks on result:

other: 365 d; anaerobic

Parent/product:

parent

Soil No.:

#1

% Degr.:

99.5

Parameter:

radiochem. meas.

Sampling time:

212 d

Remarks on result:

other: 10.23 mg/kg; aerobic

Parent/product:

parent

Soil No.:

#1

% Degr.:

95.8

Parameter:

radiochem. meas.

Sampling time:

59 d

Remarks on result:

other: 10.55 mg/kg; aerobic

Key result

Soil No.:

#1

DT50:

8.7 d

Type:

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

Temp.:

25 °C

Remarks on result:

other: aerobic; calculated outside of the study

Soil No.:

#1

DT50:

8 d

Type:

other: linear regression analysis

Temp.:

25 °C

Remarks on result:

other: aerobic; reported in original study (Spare 1995)

Soil No.:

#1

DT50:

82 d

Type:

other: linear regression analysis

Temp.:

25 °C

Remarks on result:

other: anaerobic; reported in original study (Spare 1995)

Transformation products:

not specified

Remarks:

M2, M12, M11, M10 and M9

Evaporation of parent compound:

not specified

Volatile metabolites:

yes

Residues:

yes

Aerobic:

- Microbial activity: Microbial activity of the test soil was confirmed at the start, during and the end of aerobic incubation by determining the microbial biomass (plate count method). Soils were found to be viable throughout the incubation period.

- Recovery of radioactivity: Total recovery of radioactivity ranged from 83 to 104%. The amount of soil extractable radioactivity decreased from 104% AR on day 0 to 1.8% AR on day 212 and from 103% AR on day 0 to 21% AR on day 59 (“incubation 2”). Unextractable residues increased to a maximum of 69% AR on day 28 and was between 48 and 58% AR thereafter; during “incubation 2”, unextractable residues increased to a maximum of 48% AR on day 21 and remained at that level thereafter. CO2 was evolved from the treated soil to a maximum of 33% AR after 1 year and to 20% AR on day 59 (“incubation 2”). Volatiles were insignificant. Unextractable residues in 7/9/12 months samples consisted of fulvic and humic acids (19 - 28% AR ) and humins (20 - 31% AR).

-Degradation: The test substance degraded to 3.0% AR on day 28 and to 4.4% AR on day 44 (“incubation 2”). Following the HPLC analysis, one significant unidentified fraction (“unknown 1”) was present (max 8.3% AR on day 30 and exceeding 5% AR at two consecutive time points). This fraction was characterised as very polar (no retention in reversed phase HPLC and remaining in the origin with TLC) and consists of radioactivity incorporated in high MW components as indicated by size exclusion gel-chromatography. In addition, 5 identified (M2, M12, M11, M10 and M9) and 3 unidentified metabolites (individual fractions≤4.9% AR) were observed. TLC analysis confirmed the parent quantification and metabolite pattern.

Anaerobic:

- Microbial activity: Microbial activity of the test soil was confirmed at the start, during and the end of aerobic incubation by determining the microbial biomass (plate count method). Soils and water were found to be viable throughout the incubation period.

- Recovery of radioactivity: Total recovery of radioactivity ranged from 85 to 96% AR. The amount of soil extractable radioactivity ranged from 14 to 17% AR in the first 5 month period and was 9.1% after 9 months. Unextractable residues ranged from 42 to 67% AR (max at 6 months). During the anaerobic period, CO2 evolved from the treated soil from 20% AR (start) to 26% AR after 1 year anaerobic conditions. Volatiles were insignificant. Unextractable residues in 6/8/9/12 months samples consisted of fulvic and humic acids (14 - 31% AR ) and humins (17 - 31% AR). The test substance was present at 3.0% AR at the start of the anaerobic incubation and remained stable thereafter (1.1 – 2.9 % AR). Due to the low levels of the test substance at the start of the anaerobic incubation period, no anaerobic DT50/DT90 can be calculated. HPLC analysis of the soil extracts showed 3 identified (M2, M10, M9) and 2 unidentified metabolites (individual fractions all ≤ 1.8% AR); the water layer contained 2 unidentified metabolites (all ≤ 1.2% AR). TLC analysis confirmed the parent quantification and metabolite pattern.

Table 2. Distribution of radioactivity after incubation at 25°C of sandy loam soil treated with [phenoxyphenoxy-U-14C]-labelled test substance at 10.23 mg/kg (% AR, mean of duplicate samples)

time (d)	extractables				unextrac- tables	Volatiles (C)	CO2	mass balance
	Total (A)	MeOH:aq NH4OH	0.5% NH4OH	Test substance (B)
0	104	104	n.a.	104	0.0	n.a.	n.a.	104
3	72	57	15	57	22	0.0	2.8	97
7	52	31	21	29	30	0.0	7.8	90
14	42	32	10	32	36	0.0	12	90
21	29	20	10	13	54	0.2	14	96
28	16	5.9	10	3.0	69	0.0	20	104
45	20	8.9	11	5.3	55	0.0	21	96
59	16	7.2	8.7	3.6	56	0.0	21	93
91	12	5.2	7.0	4.3	58	0.1	24	94
120	13	2.4	10	0.7	49	0.0	28	89
150	9.6	2.1	7.5	0.7	48	0.0	26	83
212	1.8	1.8	- -	0.5	58	0.0	31	91
273	- -	- -	- -	- -	- -	0.0	31	n.a.
365	- -	- -	- -	- -	- -	0.0	33	n.a.

(A) including 5 identified (M2, M12, M11, M10 and M9) and 2 unidentified metabolites (max 2.9% AR)

(B) sum of both extracts; the 3/4/5 months 0.5% NH4OH extracts were analysed for the test substance showing 0.4% AR in the 3 month sample.

(C) values provided are for ethylene glycol. Polyurethane foam plug values are all 0.0% AR

n.a.: not applicable

- -: not analysed

 

Table 3. Distribution of radioactivity after incubation at 25°C of sandy loam soil treated with [phenoxyphenoxy-U-14C]-labelled test substance at 10.55 mg/kg (“incubation 2”) (% AR, mean of duplicate samples)

time (d)	extractables					unextrac- tables	Volatiles (C)	CO2	mass balance
	Total (A)	MeOH:aq NH4OH	0.5% NH4OH	Test substance (B)	unknown 1 (B)
0	103	103	n.a.	103	- -	n.a.	n.a.	n.a.	103
3	67	58	9.3	58	2.7	23	0.0	2.9	93
7	64	52	12	55	3.0	27	0.0	7.3	98
14	36	24	12	19	4.9	46	0.0	13	95
21	32	18	14	17	8.0	48	0.0	14	94
30	22	10	12	8.2	8.3	45	0.0	18	85
44	15	7.0	8.2	4.4	5.2	48	0.0	21	83
59	21	14	7.1	4.2	4.8	44	0.0	20	85

(A) including 5 identified (M2, M12, M11, M10 and M9) and 3 unidentified metabolites (max 4.9% AR) (in addition to the one listed in the table)

(B) sum of both extracts, except for day 0 when no 0.5% NH4OH extraction was performed. Consists of radioactivity

incorporated in high MW components.

(C) both polyurethane foam plugs and ethylene glycol

n.a.: not applicable

- - : not analysed

Table 4. Distribution of radioactivity after anaerobic incubation at 25°C of sandy loam soil treated with [phenoxyphenoxy-U-14C]-labelled test substance at 10.23 mg/kg (% AR, mean of duplicate samples) 

time (m)	extractables				unextrac- tables	Volatiles(C)	CO2	water layer (D)	mass balance
	Total (A)	MeOH:aq NH4OH	0.5% NH4OH	Test substance (B)
0 (E)	16	5.9	10	3.0	69	0.0	20	n.a.	104
1	15	5.7	8.9	1.5	59	0.0	19	1.9	94
2	15	9.4	5.8	2.9	59	0.0	20	1.7	96
3	16	6.0	10	1.4	51	0.0	22	2.2	91
4	14	6.7	6.9	1.1	48	0.0	23	2.2	87
5	17	4.1	13	1.1	42	0.0	25	1.8	86
6	n.p.	4.5	n.p.	1.2	67	0.0	22	2.1	96
8	n.p.	n.p.	n.p.	n.p.	66	0.0	25	1.2	92
9	9.1	9.1	n.p.	n.p.	52	0.0	25	1.6	87
12	n.p.	n.p.	n.p.	n.p.	58	0.0	26	1.0	85

(A) including 3 identified (M2, M10, M9) and 2 unidentified metabolites (max 1.8% AR)

(B) sum of both extracts (the 2/3/4/5 months 0.5% NH4OH extracts were analysed for the test substance showing 0.2% AR in the 2 month sample)

(C) both polyurethane foam plugs and ethylene glycol

(D) 3/4/5/6/8 months samples were analysed and contained no detectable amount of the test substance and 2 unidentified metabolites (max 1.2% AR)

(E) situation at the end of 28 days aerobic incubation

n.p.: not performed;

n.a.: not applicable

Conclusions:

In a biodegradation study in Buckeystown Soil (sandy loam), performed in accordance with EPA guideline 162-1 and 162-2, the DT50 of the test substance in aerobic incubation was determined to be 8 days, and in anaerobic incubation was calculated to be 82 days. The DT50 value in aerobic incubation was re-calculated in SFO model outside of the original study, and it was 8.7 days.

Executive summary:

The biodegradation of the [U-14C-phenoxyphenoxy]-labelled test substance has been investigated in aerobic and anaerobic studies. The studies were in accordance with EPA guideline 162-1 and 162-2, and in compliance with GLP criteria. The Buckeystownsoil (sandy loan, pH 7.8) was treated with the 14C-labelled test substance at a rate of 10.23 mg/kg soil dw (for both aerobic and anaerobic incubations). The treated soil were adjusted to 75% of moisture capacity at 1/3 bar and incubated at 25 °C in the dark for up to 1 year. The soil moisture content was maintained at the initial level. Aerobic conditions were maintained by passing humidified air through the incubation flasks for ~1h at each sampling interval or every two weeks (whichever was shorter). Outgoing air was passed through 10% KOH (2 traps), polyurethane plugs (2) and ethylene glycol (1 trap) for trapping of volatiles. The study was extended with an additional set of 50 g dry weight soil samples (not pre-incubated), fortified individually at a rate of 10.55 mg/kg (added in 50 μL acetonitrile) and incubated under the same conditions for up to 2 months (“incubation 2”).

For aerobic studies, duplicate soil samples were analysed after 0, 3, 7, 14, 21, 30, 45 days, 2, 3, 4, 5, 7, 9 and 12 months (initial incubation); 0, 3, 7, 14, 21, 30, 45, and 60 days (incubation 2) post treatment. Radioactivity in post extraction soil samples was determined by combustion/LSC (humin fraction). The test substance was quantified by 1D-TLC and HPLC and identity was confirmed by co-chromatography with reference standards. The identity of the test substance in soil extracts was also confirmed by 2D-TLC and mass spectral analysis (GC-MS). Metabolites were identified by 2D-TLC and co-chromatography with reference standards. For anaerobic studies, duplicate soil samples were analysed at 1, 2, 3, 4, 5, 6, 8, 9, and 12 months post treatment. The methods for radioactivity analysis and test substance quantification were the same as for the samples from aerobic incubation.

For aerobic studies, the total recovery of radioactivity ranged from 83 to 104%. The test substance degraded to 3.0% AR on day 28 and to 4.4% AR on day 44 (“incubation 2”).Following the HPLC analysis, one significant unidentified fraction (“unknown 1”) was present (max 8.3% AR on day 30 and exceeding 5% AR at two consecutive time points).In addition, 5 identified (M2, M12, M11, M10 and M9) and 3 unidentified metabolites (individual fractions ≤ 4.9% AR) were observed. TLC analysis confirmed the parent quantification and metabolite pattern. For anaerobic studies, the total recovery of radioactivity ranged from 85 to 96% AR. The test substance was present at 3.0% AR at the start of the anaerobic incubation and remained stable thereafter (1.1 – 2.9% AR). HPLC analysis of the soil extracts showed 3 identified (M2, M9, M10) and 2 unidentified metabolites (individual fractions all ≤ 1.8% AR); the water layer contained 2 unidentified metabolites (al l ≤ 1.2% AR). TLC analysis confirmed the parent quantification and metabolite pattern.

Based on these results, the DT50 of the test substance in aerobic incubation was determined to be 8 days, and in anaerobic incubation was calculated to be 82 days. The DT50 value in aerobic incubation was re-calculated in SFO model outside of the original study, and it was 8.7 days

Reference 3

Endpoint:

biodegradation in soil: simulation testing

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

Apr 1991 to Jul 1991

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: EPA 163-1

Deviations:

not specified

GLP compliance:

yes (incl. QA statement)

Test type:

laboratory

Radiolabelling:

yes

Oxygen conditions:

aerobic

Soil classification:

USDA (US Department of Agriculture)

Year:

1991

Soil no.:

#1

Soil type:

sandy loam

% Clay:

16.9

% Silt:

30

% Sand:

49

% Org. C:

2.4

pH:

6.4

Bulk density (g/cm³):

1.14

% Moisture content:

63.6

Soil No.:

#1

Duration:

30 d

Soil No.:

#1

Initial conc.:

0.29 mg/kg soil d.w.

Based on:

test mat.

Soil No.:

#1

Initial conc.:

0.63 mg/kg soil d.w.

Based on:

test mat.

Parameter followed for biodegradation estimation:

CO2 evolution

radiochem. meas.

Soil No.:

#1

Temp.:

20°C

Humidity:

40% of MWHC (remained within 5% of the target throughout the incubation period)

Soil No.:

#1

% Total extractable:

13

% Non extractable:

61

% CO2:

15

% Recovery:

89

Remarks on result:

other: 30 d;0.29 mg/kg

Soil No.:

#1

% Total extractable:

43

% Non extractable:

51

% CO2:

3.1

% Recovery:

96

Remarks on result:

other: 3 d; 0.63 mg/kg

Parent/product:

parent

Soil No.:

#1

% Degr.:

89

Parameter:

radiochem. meas.

Sampling time:

30 d

Remarks on result:

other: 0.29 mg/kg

Parent/product:

parent

Soil No.:

#1

% Degr.:

60

Parameter:

radiochem. meas.

Sampling time:

3 d

Remarks on result:

other: 0.63 mg/kg

Key result

Soil No.:

#1

DT50:

2.9 d

Type:

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

Temp.:

20 °C

Remarks on result:

other: recalculated outside of the study

Soil No.:

#1

DT50:

2.4 d

Type:

not specified

Temp.:

20 °C

Remarks on result:

other: reported in the original study

Transformation products:

not specified

Evaporation of parent compound:

not specified

Volatile metabolites:

not specified

Residues:

yes

- Recovery of radioactivity: Total recovery of radioactivity ranged from 87 to 100% AR. The amount of extractables continuously decreased to 13% AR at day 30. Unextractable residues gradually increased to a maximum of 64% AR at day 13. CO2was evolved from the soil to 15% AR at day 30.

- Degradation: The test substance degraded to 11% AR after 13 days and remained stable thereafter. Other radioactive fractions included TLC-origin material (max 2.1% AR) and unknowns (max 1.4% AR).

 

Table 2. Distribution of radioactivity after incubation of soil treated with [Phenoxyphenoxy-U-14C] -labelled test substance at 0.3 or 0.6 mg/kg (% of applied) (mean of duplicate samples unless otherwise indicated).

Time (days)	% of applied
	total extractables	Test substance	unextractables	CO2	mass balance
	0.29 mg/kg
0	96	94	4.1	-	100
6	27	24	59	4.0	90
13 16(B)	14 20	11 14	64 58	9.2 11	87 88
30	13	11	61	15	89
	0.63 mg/kg
1 2(A)	71 54	68 51	28 43	0.1 2.4	98 99
3	43	40	51	3.1	96

(A) Situation representative for aged residues at the start of leaching experiment.

(B) Single sample.

Conclusions:

In a biodegradation study in sandy loam soil, performed in accordance with EPA guideline 163-1, the DT50 of the test substance was determined to be 2.4 days. The DT50 was re-calculated in SFO model outside of the original study, and it was 2.9 days.

Executive summary:

The biodegradation of the [phenoxyphenoxy-U-14C]-labelled test substance has been investigated in an aerobic study. The studies were in accordance with EPA guideline 163-1, and in compliance with GLP criteria. The Dielsdorf soil (sandy loan, pH 6.4) was treated with the 14C-labelled test substance ata rate of 0.29 or 0.63 mg/kg dw and incubated in the dark at 20°C for up to 30 days. The initial soil moisture content was adjusted to 40% of MWHC and remained within 5% of the target throughout the incubation period. Aerobic conditions were maintained by passing humidified, CO2free air through the incubation flasks. CO2and volatiles were trapped in ethylene glycol and 2N NaOH. Duplicate samples were analysed on days 0, 1, 2, 3, 6, 13, 16 and 30 post treatment. Radioactivity in the extracts and liquid traps was determined by LSC. Radioactivity in post-extraction solids was determined by combustion/LSC. The acetonitrile soil extract was concentrated prior to TLC analysis. Compound identification was by co-chromatography with unlabelled reference standards.

Total recovery of radioactivity ranged from 87 to 100% AR. The amount of extractables continuously decreased to 13% AR at day 30. Unextractable residues gradually increased to a maximum of 64% AR at day 13. CO2 was evolved from the soil to 15% AR at day 30. The test substance degraded to 11% AR after 13 days and remained stable thereafter. Other radioactive fractions included TLC-origin material (max 2.1% AR) and unknowns (max 1.4% AR). Based on these results. the DT50 of the test substance was determined to be 2.4 days. The DT50 was re-calculated in SFO model outside of the original study, and it was 2.9 days.

Reference 4

Endpoint:

biodegradation in soil: simulation testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 Feb 2009 to 31 Mar 2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 307 (Aerobic and Anaerobic Transformation in Soil)

Version / remarks:

April 2002

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

laboratory

Radiolabelling:

no

Oxygen conditions:

aerobic

Soil classification:

USDA (US Department of Agriculture)

Year:

2009

Soil no.:

#1

Soil type:

sandy clay loam

% Clay:

25

% Silt:

24

% Sand:

51

% Org. C:

2.4

pH:

6.2

CEC:

18.9 meq/100 g soil d.w.

Bulk density (g/cm³):

1.5

Details on soil characteristics:

An overview of the properties of the soil is provided in Table 1 in 'Any other information on materials and methods incl. tables'.
SOIL COLLECTION AND STORAGE
- Geographic location: Jealott's Hill Farm, Nupton Road, Warfield, Bracknell, UK
- Date of field sampling: 29 January 2009
- Sampling depth (cm): 5 - 15 cm
- Storage conditions: Prior to use the soil was stored in the dark, in an incubator routinely maintained at 4 ± 2 ˚C, in loosely tied plastic bags in accordance with ISO 10381-6.
- Soil preparation: Thoroughly mixed and passed through a 2 mm mesh sieve with the minimum of air drying.

PROPERTIES OF THE SOILS (in addition to defined fields)
- Water Holding capacity at pF 0 MWHC (0.001 bar): 64.5%
- Water Holding capacity at pF 2.0 (0.1 bar): 29.8%
- Water Holding capacity at pF 2.5 (0.33 bar): 23.6%
- Microbial biomass: 460.0 µg C/g
- Microbial biomass: 1.9% of OC

Soil No.:

#1

Duration:

10 d

Soil No.:

#1

Initial conc.:

0.3 mg/kg soil d.w.

Based on:

act. ingr.

Parameter followed for biodegradation estimation:

test mat. analysis

Soil No.:

#1

Temp.:

20 ± 2°C

Humidity:

approximately pF2

Microbial biomass:

460.0 µg C/g

Details on experimental conditions:

An overview of the study design is provided in Table 2 in 'Any other information on materials and methods incl. tables'.
EXPERIMENTAL DESIGN
- Soil preincubation conditions: The soil was pre-incubated in the soil vessels under study conditions for 7 days prior to treatment with the application solution.
- Soil (g/replicate): 50 g (dry weight) per replicate
- Soil condition: Fresh, passed through 2 mm sieve prior to use
- No. of replication treatments: 2
- Test apparatus: Plastic Incubation Vessels
- Evaporation of application solvent: Yes

Test material application
- Application method: Aliquots of the test substance stock solution (150 μg/mL) were used to dose the main study units. The test substance was applied to 18 Acres soil on 10 March 2009. The treatment solution was applied to each soil vessel dropwise in acetonitrile (100 μL). An aliquot (133 μL) of the treatment solution was transferred to a 20 mL volumetric flask on the day of application. The volume in each flask was made up to the mark with methanol: water (40/60 v/v) giving a test substance concentration of 1 μg/mL. An aliquot (50 μL) of the 1 μg/mL solution was transferred to a 25 mL volumetric flask and made up to the mark with methanol: water (40/60 v/v) giving a test substance concentration of 0.002 μg/mL. Triplicate aliquots of the 0.002 μg/mL solution were quantified by LC-MS/MS to confirm the concentration of the treatment solution.
- Volume of test solution used/treatment: 100 μL
- Any indication of test material adsorbing to walls of test apparatus: No
- Moisture maintenance method: The moisture content of the soils was checked by weighing the vessels at regular intervals of up to eight days. Any moisture losses were replaced on these occasions by the addition of reverse osmosis water.
- Continuous darkness: Yes

SAMPLING DETAILS
- Sampling intervals: Soil vessels were extracted in duplicate at each sampling time. Samples were taken immediately after treatment (zero time) and at 1, 3, 7 and 10 DAT. Extraction of soils commenced on the sampling dates.

Soil No.:

#1

% Recovery:

95.9

Remarks on result:

other: on day 0

Parent/product:

parent

Soil No.:

#1

% Degr.:

> 95

Parameter:

test mat. analysis

Sampling time:

10 d

Key result

Soil No.:

#1

DT50:

1.5 d

Type:

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

Temp.:

20 °C

Transformation products:

not measured

Evaporation of parent compound:

not specified

Volatile metabolites:

not specified

Residues:

not specified

Details on results:

An overview of the results is provided in Table 3 - Table 4 in 'Any other information on results incl. tables'.
- Recovery: Mean recoveries of the test substance obtained from control soil systems fortified at 0.015 and 0.3 mg/kg ranged from 90 to 97%, with an overall mean recovery of 94%. Overall mean recovery was within the acceptable range of 70 to 110%. The method performance was therefore considered appropriate for use within this study.
- Limit of Quantification (LOQ): Acceptable mean recovery (70 to 110%), and a relative standard deviation (RSD) of ≤ 20% were obtained for the analysis of the test substance at the lowest fortification level of 0.015 mg/kg. Therefore, the limit of quantification established for the supplied method of 0.01 mg/kg was accepted.
- Quantification of Degradation: The analytical methods employed were performing at an overall mean recovery efficiency of 94% during this study. Percentage of dosed test substance values have been corrected for any apparent residue measured in the corresponding control soil extracted at the time of analysis, where appropriate.
- DegT50 of the test substance in soil: Because the concentrations of the test substance in the 10 DAT samples were below 5% of applied test article, the study was terminated early with the 20 and 30 DAT timepoints not analysed. The single first-order DegT50 value for the degradation of the test substance in 18 Acres soil was 1.5 days.

Table 3. Percent Recovery of applied test substance from Soil

Sampling Interval (DAT)	0	1	3	7	10
Unit Code	A01   A02	A03   A04	A05   A06	A07  A08	A09   A10
Total in soil	97.0   94.8	54.6  53.4	30.9  18.8	7.3    7.2	3.8    4.0
Mean	95.9	54.0	24.9	7.3	3.9

 

Table 4. Concentration of the test substance from Soil (mg/kg) 

Sampling Interval (DAT)	0		1		3		7		10
Unit Code	A01	A02	A03	A04	A05	A06	A07	A08	A09	A10
Total in soil	0.2909	0.2843	0.1637	0.1603	0.0928	0.0564	0.0220	0.0216	0.0115	0.0120
Mean	0.2876		0.1620		0.0746		0.0218		0.0118

 

Conclusions:

In a biodegradation study in soil, performed in accordance with OECD TG 307, the calculated DT50 values using single first-order kinetics was 1.5 days (18 Acres soil).

Executive summary:

 The biodegradation of the test substance has been investigated under aerobic conditions at 20 ± 2 °C in one soil, 18 Acres (sandy clay loam, pH 6.2). The study was performed in accordance with OECD TG 307, and in compliance with GLP criteria. The test substance was applied to the surface of individual soil samples at a nominal application rate of 0.3 mg/kg soil dw. The soil was incubated under aerobic conditions in the laboratory and maintained under moist (approximately pF2), dark conditions for up to 10 days. Duplicate samples were taken for analysis at 0, 1, 3, 7 and 10 days after treatment (DAT). At each sampling time, the soil samples were extracted using suitable solvents and the test substance present in the extracts was quantified by LC-MS/MS. The methodology was based upon a validated soil residue method. 

The total amount of the test substance in the soil extracts declined steadily over the 10 day duration of the study in both soils, from 95.9% (DAT 0) to 3.9% (DAT 10).Therefore,the study was terminated early with the 20 and 30 DAT timepoints not analysed. The single first-order DT50 value for the degradation of the test substance in 18 Acres soil was 1.5 days.

 

 

Reference 5

Endpoint:

biodegradation in soil: simulation testing

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

01 Feb 2001 to 29 Jun 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 307 (Aerobic and Anaerobic Transformation in Soil)

Version / remarks:

Draft OECD Guideline for Testing of Chemicals, Aerobic and Anaerobic Transformation in Soil, July 1999

Deviations:

not specified

Qualifier:

according to guideline

Guideline:

other: Dutch Registration Guideline G.1

Version / remarks:

Behaviour in soil; Ministry of Agriculture and Fisheries, Ministry of Public Health and
Environmental Hygiene, Ministry of Social Affairs, January 1987

Deviations:

not specified

GLP compliance:

yes (incl. QA statement)

Test type:

laboratory

Radiolabelling:

yes

Oxygen conditions:

aerobic

Soil classification:

USDA (US Department of Agriculture)

Year:

2000

Soil no.:

#1

Soil type:

silt loam

% Clay:

11.8

% Silt:

52.3

% Sand:

35.9

% Org. C:

2.1

pH:

7.3

CEC:

13.9 meq/100 g soil d.w.

Soil No.:

#1

Duration:

120 d

Soil No.:

#1

Initial conc.:

0.64 mg/kg soil d.w.

Based on:

test mat.

Remarks:

corresponding to a field application rate of about 420 g a.i./ha

Parameter followed for biodegradation estimation:

CO2 evolution

radiochem. meas.

Soil No.:

#1

Temp.:

9.1 ± 0.4 °C

Humidity:

40% WHC

Microbial biomass:

55.0 mg microbial carbon per 100 g dry soil (on day 0)

Soil No.:

#1

Temp.:

19.8 ± 0.5 °C

Humidity:

40% WHC

Microbial biomass:

55.0 mg microbial carbon per 100 g dry soil (on day 0)

Soil No.:

#1

Temp.:

19.8 ± 0.5 °C

Humidity:

25% WHC

Microbial biomass:

55.0 mg microbial carbon per 100 g dry soil (on day 0)

Soil No.:

#1

Temp.:

30.3 ± 0.1 °C

Humidity:

40% WHC

Microbial biomass:

55.0 mg microbial carbon per 100 g dry soil (on day 0)

Soil No.:

#1

% Total extractable:

2

% Non extractable:

60.5

% CO2:

36.4

% Other volatiles:

< 0.1

% Recovery:

99

Remarks on result:

other: 20 °C and 40% WHC; on day 120

Soil No.:

#1

% Total extractable:

2.7

% Non extractable:

62.1

% CO2:

31.3

% Other volatiles:

< 0.1

% Recovery:

96.1

Remarks on result:

other: 20 °C and 25% WHC; on day 120

Soil No.:

#1

% Total extractable:

3.5

% Non extractable:

64.3

% CO2:

27.2

% Other volatiles:

< 0.1

% Recovery:

95

Remarks on result:

other: 10 °C and 40% WHC; on day 120

Soil No.:

#1

% Total extractable:

1.8

% Non extractable:

50

% CO2:

46.3

% Other volatiles:

< 0.1

% Recovery:

98.1

Remarks on result:

other: 30 °C and 40% WHC; on day 120

Parent/product:

parent

Soil No.:

#1

% Degr.:

98.6

Parameter:

radiochem. meas.

Sampling time:

28 d

Remarks on result:

other: 20 °C and 40% WHC

Parent/product:

parent

Soil No.:

#1

% Degr.:

97.7

Parameter:

radiochem. meas.

Sampling time:

56 d

Remarks on result:

other: 20 °C and 25% WHC

Parent/product:

parent

Soil No.:

#1

% Degr.:

96.6

Parameter:

radiochem. meas.

Sampling time:

120 d

Remarks on result:

other: 10 °C and 40% WHC

Parent/product:

parent

Soil No.:

#1

% Degr.:

98.3

Parameter:

radiochem. meas.

Sampling time:

28 d

Remarks on result:

other: 30 °C and 40% WHC

Key result

Soil No.:

#1

DT50:

1.3 d

Type:

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

Temp.:

20 °C

Remarks on result:

other: 20 °C and 40% WHC

Soil No.:

#1

DT50:

2.7 d

Type:

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

Temp.:

20 °C

Remarks on result:

other: 20 °C and 25% WHC

Soil No.:

#1

DT50:

4.5 d

Type:

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

Temp.:

20 °C

Remarks on result:

other: 10 °C and 40% WHC

Soil No.:

#1

DT50:

1.1 d

Type:

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

Temp.:

20 °C

Remarks on result:

other: 30 °C and 40% WHC

Transformation products:

not specified

Remarks:

M2

Evaporation of parent compound:

not specified

Volatile metabolites:

yes

Residues:

yes

Results

Microbial activity of the test soil was confirmed at the start and the end of aerobic incubation by determining the microbial biomass. Total recovery of radioactivity ranged from 92 to 106% AR. The amount of soil extractable radioactivity decreased from 101 - 105% AR on day 0 to 1.8 - 3.5% AR on day 120. Unextractable residues increased from 0.2 - 0.4% AR on day 0 to 70% AR on day 28/56/14 at 9°C/20°C (25% MWHC)/20°C (40% MWHC) and to 66% AR on day 14 at 30°C, showing a decline to 64/62/61/50% AR at 9°C/20°C (25% MWHC)/20°C (40% MWHC)/30°C on day 120. CO2 was evolved from the treated soils to a maximum of 27-46% AR on day 120. Volatile organic compounds were insignificant. The test substance degraded to 3.1% AR after 14 days during incubation at 40% MWHC and 20°C. Degradation at 25% MWHC (to 3.9% AR after 28 days) and 9°C (to 4.4% AR after 28 days) was slower, at 30°C faster (to 3.1% AR after 7 days). Seven unidentified metabolites and one identified (M2) metabolite were found in low amounts (≤ 4.5% AR). TLC-origin material was ≤ 5.3% AR. Further extraction of day 3 and day 90 samples released 0.7 - 4.0% AR. Unextractable soil residues in the 90 day samples were shown to consist of humic acids (up to 5.9% AR), humin (up to 58% AR) and fulvic acids (up to 2.3% AR).

Table 2. Distribution of Radioactivity in Gartenacker Soil incubated at 20 °C and 40% WHC (Series 1)

Incubation time (day)	Replicate	Cold Extract	Reflux	Sum	14CO2	Organic volatiles	Non-Extractable	Recovery
	% of applied radioactivity
0	1	104	0.5	104.5	-	-	0.5	105.0
0	2	104.9	0.6	105.5	-	-	0.4	105.8
0	mean	104.4	0.6	105	-	-	0.4	105.4
3	1	25.2	0.9	26.1	10.9	<0.1	55.6	92.7
7	1	9.8	0.8	10.6	17.4	<0.1	66.6	94.7
7	2	10	0.8	10.8	17.4	<0.1	65.2	93.4
7	mean	9.9	0.8	10.7	17.4	<0.1	65.9	94.0
14	1	4.5	0.6	5.1	23.00	<0.1	69.8	97.9
28	1	3.5	0.4	3.9	27.6	<0.1	66.9	98.4
28	2	2.6	0.5	3.1	27.8	<0.1	66.4	97.3
28	mean	3.1	0.4	3.5	27.7	<0.1	66.6	97.8
56	1	2.1	0.6	2.6	29.3	<0.1	68.7	100.7
56	2	2.2	0.5	2.7	29.6	<0.1	61.8	94.0
56	mean	2.1	0.5	2.7	29.5	<0.1	65.3	97.4
90	1	2.00	0.6	2.5	31.5	<0.1	62.6	96.6
120	1	1.7	0.4	2.00	36.0	<0.1	61.8	99.9
120	2	1.6	0.4	2.00	36.8	<0.1	59.3	98.1
120	mean	1.6	0.4	2.00	36.4	<0.1	60.5	99.0

  

Table 3. Distribution of Radioactivity in Gartenacker Soil incubated at 20 °C and 25% WHC (Series 2)

Incubation time (day)	Replicate	Cold Extract	Reflux	Sum	14CO2	Organic volatiles	Non-Extractable	Recovery
	% of applied radioactivity
0	1	101.7	0.6	102.3	-	-	0.4	102.7
0	2	105.0	0.6	105.6	-	-	0.4	106.0
0	mean	103.4	0.6	103.9	-	-	0.4	104.3
3	1	49.0	0.8	49.8	7.0	<0.1	40.2	97.0
7	1	24.1	1.0	25.2	14.6	<0.1	58.8	98.6
7	2	27.8	0.9	28.7	14.6	<0.1	55.5	98.9
7	mean	26.0	1.0	26.9	14.6	<0.1	57.1	98.7
14	1	14.4	0.7	15.1	13.8	<0.1	63.5	92.4
28	1	5.9	0.6	6.5	24.0	<0.1	68.5	98.9
28	2	5.3	0.5	5.8	24.0	<0.1	68	97.8
28	mean	5.6	0.6	6.1	24.0	<0.1	68.3	98.4
56	1	3.0	0.6	3.7	27.0	<0.1	71	101.7
56	2	2.9	0.7	3.6	27.0	<0.1	69.2	99.8
56	mean	3.0	0.7	3.6	27.0	<0.1	70.1	100.7
90	1	2.4	0.5	2.9	31.7	<0.1	63.4	98.0
120	1	2.1	0.6	2.7	31.3	<0.1	62.9	96.9
120	2	2.1	0.6	2.7	31.4	<0.1	61.3	95.4
120	mean	2.1	0.6	2.7	31.3	<0.1	62.1	96.1

Table 4. Distribution of Radioactivity in Gartenacker Soil incubated at 10 °C and 40% WHC (Series 3)

Incubation time (day)	Replicate	Cold Extract	Reflux	Sum	14CO2	Organic volatiles	Non-Extractable	Recovery
	% of applied radioactivity
0	1	102.7	0.5	103.2	-	-	0.3	103.4
0	2	101.7	0.4	102.1	-	-	0.3	102.4
0	mean	102.2	0.4	102.6	-	-	0.3	102.9
3	1	66.5	0.8	67.4	1.9	<0.1	25.7	94.9
7	1	39.0	0.9	39.8	5.6	<0.1	48.7	94.2
7	2	46.1	1.0	47.1	5.6	<0.1	43.7	96.4
7	mean	42.5	0.9	43.5	5.6	<0.1	46.2	95.3
14	1	14.9	0.8	15.7	15.1	<0.1	64.7	95.5
28	1	7.1	0.7	7.7	22.1	<0.1	68.8	98.6
28	2	6.8	0.5	7.3	22.3	<0.1	70.6	100.3
28	mean	6.9	0.6	7.5	22.2	<0.1	69.7	99.5
56	1	3.8	0.8	4.7	23.5	1.1	70.8	100.0
56	2	3.9	0.8	4.7	23.5	1.1	60.7	90.0
56	mean	3.9	0.8	4.7	23.5	1.1	65.7	95.0
90	1	3.3	0.7	3.9	23.5	1.1	67.9	101.2
120	1	2.9	0.6	3.5	27.1	<0.1	65.4	96.1
120	2	3.0	0.6	3.6	27.2	<0.1	63.2	94.0
120	mean	2.9	0.6	3.5	27.2	<0.1	64.3	95.0

Table 5. Distribution of Radioactivity in Gartenacker Soil incubated at 30 °C and 40% WHC (Series 4)

Incubation time (day)	Replicate	Cold Extract	Reflux	Sum	14CO2	Oragnic volatiles	Non-Extractable	Recovery
	% of applied radioactivity
0	1	100.8	0.5	101.3	-	-	0.2	101.6
0	2	100.4	0.6	101.0	-	-	0.2	101.2
0	mean	100.6	0.6	101.2	-	-	0.2	101.4
3	1	25.0	0.9	25.9	13.8	<0.1	52.7	92.4
7	1	6.7	0.7	7.4	20.4	<0.1	63.8	91.6
7	2	8.1	0.8	8.8	20.4	<0.1	63.1	92.3
7	mean	7.4	0.8	8.1	20.4	<0.1	63.5	92.0
14	1	5.0	0.6	5.7	29.1	<0.1	65.5	100.3
28	1	2.5	0.5	3.0	35.0	<0.1	61.8	99.8
28	2	2.8	0.5	3.3	34.7	<0.1	56.7	94.6
28	mean	2.7	0.5	3.1	34.8	<0.1	59.2	97.2
56	1	1.7	0.4	2.1	40.3	<0.1	62.0	104.4
56	2	1.7	0.5	2.3	40.1	<0.1	65.7	108.0
56	mean	1.7	0.5	2.2	40.2	<0.1	63.8	106.2
90	1	1.5	0.5	2.0	44.3	<0.1	53.8	100.0
120	1	1.5	0.4	1.9	46.5	<0.1	52.2	100.6
120	2	1.2	0.4	1.7	46.1	<0.1	47.8	95.6
120	mean	1.4	0.4	1.8	46.3	<0.1	50.0	98.1

 

Table 6. Distribution of the test substance and Metabolites in Gartenacker Soil incubated at 20 °C and 40% WHC (Series 1)

Incubation time (day)	Replicate	Parent	Origin	U1	U3	U4	M2	U7	U8	U10
	% of applied radioactivity
0	1	104.5	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
0	2	105.5	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
0	mean	105.0	<LD	l<LD	<LD	<LD	<LD	<LD	<LD	<LD
3	1	18.1	4.4	2.3	<LD	1.4	<LD	<LD	<LD	<LD
7	1	7.8	2.7	<LQ	<LD	<LD	<LD	<LD	<LD	<LD
7	2	7.0	2.3	1.5	<LD	<LD	<LD	<LD	<LD	<LD
7	mean	7.4	2.5	0.8	<LD	<LD	<LD	<LD	<LD	<LD
14	1	3.1	1.5	<LD	0.1	0.1	0.2	0.1	0.1	<LD
28	1	1.5	2.4	<LD	<LD	<LD	<LD	<LD	<LD	<LD
28	2	1.3	0.9	<LD	<LQ	0.1	<LQ	0.1	<LD	0.8
28	mean	1.4	1.6	<LD	<LD	< 0.1	<LD	< 0.1	<LD	0.4
56	1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
56	2	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
56	mean	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
90	1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	2	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	mean	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD

 

Table 7. Distribution of the test substance and Metabolites in Gartenacker Soil incubated at 20 °C and 25% WHC (Series 2)

Incubation time (day)	Replicate	Parent	Origin	U1	U3	U4	M2	U7	U10	U11
	% of applied radioactivity
0	1	102.3	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
0	2	105.6	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
0	mean	103.9	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
3	1	41.4	3.6	2.0	0.5	1.1	<LQ	<LD	<LD	<LD
7	1	18.2	2.3	3.3	<LD	<LD	<LQ	<LD	<LD	<LQ
7	2	20.7	2.6	3.8	<LD	<LD	<LQ	<LD	<LD	<LQ
7	mean	19.5	2.4	3.5	<LD	<LD	<LQ	<LD	<LD	<LQ
14	1	10.8	2.2	1	0.1	0.1	0.3	0.1	0.1	0.3
28	1	4.4	1.7	<LQ	<LD	<LD	<LQ	<LD	<LD	<LD
28	2	3.3	2.1	<LQ	<LD	<LD	<LQ	<LD	<LD	<LD
28	mean	3.9	1.9	<LQ	<LD	<LD	<LQ	<LD	<LD	<LD
56	1	2.4	1.1	<LQ	<LQ	<LQ	<LQ	<LQ	<LD	<LD
56	2	2.2	1.3	<LD	<LD	<LD	<LD	<LQ	<LD	<LD
56	mean	2.3	1.2	<LD	<LD	<LD	<LD	<LQ	<LD	<LD
90	1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	2	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	mean	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD

 

Table 8. Distribution of the test substance and Metabolites in Gartenacker Soil incubated at 10 °C and 40% WHO (Series 3)

Incubation time (day)	Replicate	Parent	Origin	U1	U3	U4	M2	U7	U8	U10
	% of applied radioactivity
0	1	103.2	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
0	2	102.1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
0	mean	102.6	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
3	1	59.0	5.3	1.0	1.1	0.9	<LD	<LD	<LD	<LD
7	1	31.2	3.2	2.3	1.0	2.1	<LD	<LD	<LD	<LD
7	2	40.4	2.0	1.7	1.3	1.6	<LD	<LD	<LD	<LD
7	mean	35.8	2.6	2.0	1.2	1.8	<LD	<LD	<LD	<LD
14	1	11.1	2.6	1.1	<LQ	0.4	0.2	<LQ	<LD	<LQ
28	1	4.3	2.4	0.4	<LQ	0.2	0.2	<LD	<LD	<LQ
28	2	4.4	2.2	0.3	0.1	<LD	0.4	<LD	<LD	<LD
28	mean	4.4	2.3	0.4	0.1	0.1	0.3	<LD	<LD	<LD
56	1	2.7	1.7	<LD	<LD	<LD	<LD	0.1	<LD	<LD
56	2	3.4	1.3	<LD	<LD	<LD	<LD	<LD	<LD	<LD
56	mean	3.0	1.5	<LD	<LD	<LD	<LD	<LD	<LD	<LD
90	1	2.9	1.0	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	1	120
120	2	3.4	0.1	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	mean	3.4	0.2	<LD	<LD	<LD	<LD	<LD	<LD	<LD

 

Table 9. Distribution of the test substance and Metabolites in Gartenacker Soil incubated at 30 °C and 40% WHC (Series 4)

Incubation time (day)	Replicate	Parent	Origin	U1	U3	U4	M2	U11	U8	U10
	% of applied radioactivity
0	1	101.3	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
0	2	101.0	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
0	mean	101.2	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
3	1	15.5	4.1	4.5	<LQ	<LD	<LD	1.3	<LD	<LD
7	1	3.2	3.7	<LD	<LD	<LD	<LD	<LD	<LD	<LD
7	2	3.0	5.9	<LD	<LD	<LD	<LD	<LD	<LD	<LD
7	mean	3.1	4.8	<LD	<LD	<LD	<LD	<LD	<LD	<LD
14	1	3.7	1.5	0.1	<LQ	<0.1	0.1	<LQ	<LQ	<LQ
28	1	1.9	0.9	<LD	<LD	<LD	<LQ	<LD	<LD	<LD
28	2	1.5	1.8	<LD	<LD	<LD	<LD	<LD	<LD	<LD
28	mean	1.7	1.3	<LD	<LD	<LD	<LD	<LD	<LD	<LD
56	1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
56	2	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
56	mean	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
90	1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	1	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	2	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD
120	mean	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD

 

 Table 10. Degradation of the Test Substance

	Series 1 20 °C 40% WHC	Series 2 20 °C 25% WHC	Series 3 10 °C 40% WHC	Series 4 30 °C 40% WHC
Half-life(d)	1.3	2.7	4.5	1.1
DT90 (d)	4.3	9.0	14.8	3.8

Conclusions:

In a biodegradation study in Gartenacker Soil, performed in accordance with OECD TG 307 draft, the test substance was very rapidly degraded at 30 °C and 20 °C with a DT50 of 1.1 to 1.3 days. Decreasing the temperature to 10 °C resulted in an increase of the DT50 by a factor of approximately 3.5 (4.5 days). When the soil moisture content was lowered to 25 % WHC at 20°C, the DT50 of the test substance was 2.7 days.

Executive summary:

The biodegradation of the [phenoxy-U-14C]-labelled test substance has been investigated under aerobic conditions in Gartenacker soil (loam/silt loam, pH 7.3). The study was performed in accordance with the draft of OECD TG 307 and the Dutch national guideline G1, and in compliance with GLP criteria. The soil was treated with the 14C-labelled test substance at a concentration of 0.64 mg/kg dw soil (corresponding to a field rate of 420 g active ingredient/ha). The soils were incubated under aerobic conditions in the laboratory and maintained under moist in the dark 20 °C with a soil moisture content of 40% maximum water holding capacity (WHC; series 1) and of 25% WHC (series 2). In addition soil samples were incubated with a soil moisture of 40% WHC at 10 °C (series 3) and at 30 °C (series 4) for up to 120 days. For each soil, duplicate samples were taken for analysis at 0, 3, 7, 14, 28, 56, 90 and 120 days after treatment (DAT). At each sampling time, the soil samples were extracted using suitable solvents and the test substance present in the extracts was quantified by TLC and HPLC. The amount of evolved 14CO2 was measured by LSC.

The overall recovery comprising the soil extracts, non-extractable residues and volatile products for all series was between 90.0% and 108.0%. The amount of the test substance declined from 105.0% on day 0 to 1.4% on day 28 (series 1), from 103.9% to 2.3% (series 2) on day 56, from 102.6% to 3.4% on day 120 (series 3) and from 101.2% to 1.7 %on day 28 (series 4). Based on chromatographic analysis besides the parent molecule several minor metabolites, were found in amounts 5.3%. One of them was identified as M2, by co-chromatography using 2D-TLC analysis. In conclusion, the test substance was very rapidly degraded at 30 °C and 20 °C with a half-life of 1.1 to 1.3 days. Decreasing the temperature to 10 °C resulted in an increase of the half-life by a factor of approximately 3.5 (4.5 days). When the soil moisture content was lowered to 25 % WHC at 20°C, the half-life of the test substance was 2.7 days. Under all conditions, the test substance was mainly degraded by mineralisation and formation of bound residues. The amount of non-extractable residues at study end ranged from 50.0 % to 64.3 %. Carbon dioxide accounted for up to 46.3 %.

Description of key information

All available data was assessed and the aerobic studies following standard test guidelines are included here in a weight of evidence approach.

Geometric mean DT50 in soil = 3.5 d, 20 °C, OECD TG 307, OECD TG 307 draft, EPA 162 - 1, EPA 162 - 2 and (or) EPA 163 -1 guidelines followed studies, Brice 2009, Nicollier 2001, Spare and Thede 1995a, Spare and Thede 1995b, and Rümbeli 1991.

Key value for chemical safety assessment

Half-life in soil:

3.5 d

at the temperature of:

20 °C

Additional information

Table. DT50 values for the test substance in soil under aerobic test conditions

USDA / Name / Origin	OC [%] / pH (water)	T. [°C] / Moisture	DT50 [d] - Kinetic model in original report	DT50 [d] - Kinetic model recalculated	Author / Year
Silt loam, Les Barges, CH	2.1 / 7.3 (KCl)	20 / 40% MWHC	1.3*- SFO	1.3* - SFO	Nicollier & Adam 2001
Sandy loam, Lyme Kiln, Maryland, USA	1.3 / 7.7 Phenyl-A label	25 / 75% FC	0.28 – primary 237 -secondary	81.5 - SFO	Spare, 1995, Thede, 1995
Sandy loam, Lyme Kiln, Maryland, USA	1.3 / 7.8 Phenyl-B label	25 / 75% FC	8 – primary 82 – secondary	8.7 - SFO	Spare, 1995, Thede, 1995
Geomean for Lyme Kiln				26.6* - SFO
Sandy loam, Dielsdorf, CH	2.4 / 6.4	20 / 40% MWHC	2.4	2.9* - SFO	Rümbeli, 1991
Sandy clay loam, 18 Acres, UK	2.4 / 7.2	20 / pF 2	1.5* – SFO	1.5* - SFO	Brice, 2009
Geomean (n=4)				3.5

*Value used for determination of the geomean

Soil dissipation

A large number of soil dissipation studies were conducted with the test material. As information on soil dissipation is outside the scope of REACH, the studies are not summarized as an endpoint study entry but briefly discussed here. The test material was tested for different applications in different crops and bare grounds with a rate ranging between 11 - 1369 g/ha. The DT50 reported in the original reports as calculated with (linear) SFO model were between 4.0 - 55 days. The DT50 as recalculated SFO model were between 2.3 - 37.5 days. The geometric mean DT50 (n=17) for soil dissipation was calculated to be 8.1 days (Hänni 1990; Dorn 2003; Schwager 1990; Andre 1991; Emburey 2004; Mc Donald 1995; Obsirst 1995; Jacobson 1995; Schuster 1995; Rice 1995; Fyler 1983 and Fyler 1985 a,b;).