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Biodegradation in water and sediment: simulation tests

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Endpoint:
biodegradation in water and sediment: simulation testing, other
Remarks:
natural water/sediment systems (pond and river).
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods
Qualifier:
according to
Guideline:
other: Procedures for Assessing the Environmental Fate and Ecotoxicity of Pesticides, 8.2. Aerobic Aquatic Degradation, Society of Environmental Toxicology and Chemistry, SETAC Europe.
Qualifier:
according to
Guideline:
other: Richtlinie für die Prüfung von Pflanzenschutzmitteln, Teil IV, 5-1, Abbaubarkeit und Verbleib von Pflanzenschutzmitteln im Wasser/Sediment-System. Biologische Bundesanstalt für Land- und Forstwirtschaft Bundesrepublik Deutschland
Version / remarks:
December 1990
GLP compliance:
yes
Radiolabelling:
yes
Remarks:
14C-labelled at position 2 of thiazole ring
Oxygen conditions:
aerobic
Inoculum or test system:
natural water / sediment: freshwater
Details on source and properties of surface water:
SOURCE
- River water: Rhine River at Möhlin (AG, Switzerland)
- Pond water: Fröschweiher at Rheinfelden (AG, Switzerland)

SAMPLING
- Water was sampled down to a depth of 5 to 10 cm with a plastic container. The sampling site was located about 5 m from the shore. Water samples were sieved (0.2 mm) and transported to the laboratory in sealed plastic containers.

PROPERTIES OF RIVER WATER
- Oxygen: 10.2 mg/L at the surface; 9.4 mg/L at 5 cm above sediment
- pH: 8.25 at the surface and at 5 cm above sediment.
- Redox potential: 295 mV at the start of acclimation

PROPERTIES OF POND WATER
- pH: 7.86 at the surface, 7.87 at 5 cm over sediment
- Oxygen: 0.9 mg/L at the surface, 0.86 mg/L 5 cm over sediment
- Redox potential: 282 mV at the start of acclimation
Details on source and properties of sediment:
SOURCE
- River sediment: Same as water
- Pond sediment: Same as water

SAMPLING
Sediment was sampled with a shovel from the top 5 to 10 cm of each system. The sampling site was located about 5 m from the shore. Sediments were sieved (2 mm) and transported to the laboratory in sealed plastic containers.

PROPERTIES OF RIVER SEDIMENT
- % Clay (<2 µm): 27.8
- % Silt (50-2 µm): 41.3
- % Sand (2000-50 µm: 30.9
- Soil type (USDA): Not reported (but loam, based on reported particle size distribution)
- pH: 7.5
- Redox potential (mV): -473 at start of acclimation
- Organic matter (%): 2.9
- Organic carbon (%): 1.7
- Cation Exchange Capacity (mmol/z/100g sediment): 10.9
- Nitrogen (total, %): 0.14
- Phosphorus (total, mg/kg): 619
- Maximum water capacity, % (0.01 bar): 56.10

PROPERTIES OF POND SEDIMENT
- % Clay (<2 µm): 9.8
- % Silt (50-2 µm): 48.4
- % Sand (2000-50 µm): 41.8
- Soil type (USDA): Not reported (but clay loam, based on reported particle size distribution)
- pH: Not reported
- Redox potential (mV): -437 at start of acclimation
- Organic matter (%): 3.8
- Organic carbon (%): 2.2
- Cation Exchange Capacity (mmol/z/100g sediment): 17.8
- Nitrogen (total, %): 0.19
- Phosphorus (total, mg/kg): 768
- Maximum water capacity, % (0.01 bar): 50.9
Duration of test (contact time):
100 d
Initial conc.:
0.104 mg/L
Based on:
test mat.
Remarks:
reported as 1.5 times the field application rate assuming a homogeneous distribution of the compound within the water layer down to a depth of 30 cm
Parameter followed for biodegradation estimation:
CO2 evolution
radiochem. meas.
Details on study design:
TEST SYSTEM
- Apparatus: 1L incubation flasks (internal diameter 10 cm), in the dark.
- Number of replicates: Duplicate samples of each water/sediment system for each sampling interval; high dose samples: eight in total
- Sediment condition: Passed through 2 mm sieve prior to use
- Water condition: Passed through 0.2 mm sieve prior to use and any particulates allowed to settle before use
- Control conditions: Not applicable
- Incubation conditions: Under aeration with moistened air (flow rate 60-100 mL/min). Gentle agitation of the water phase without disturbing the sediment was achieved by means of a suspended magnetic stirrer. Effluent air, from each individual incubation vessel, was passed through a series of volatile traps. Aquatic systems were acclimatised to the test conditions for a period of about 3 weeks.

SAMPLE SIZES
- River sediment (Rhine): 226.70 g wet weight (127.60 g dry weight)
- Pond sediment (Rheinfelden): 208.08 g wet weight (115.71 g dry weight)
- River water: 500 mL (about 6 cm) for both systems resulting in 4:1 water: sediment ratio)
- Pond water (Rheinfelden): 212 mL (resulting in 4:1 water: sediment ratio)

TEST MATERIAL APPLICATION
- Solvent: Acetonitrile
- Volume of test solution used per treatment: Two treatment solutions were used; treatment solution 1 for 20 flasks intended to be incubated longer than 8 days and 8 high dose samples and treatment solution 2 for 8 flasks intended to be incubated shorter than 8 days. The volumes used (to treat 500 mL water layer) were 51 µL (normal rate, 0.1 ppm), 510µL (higher rate, 1 ppm) for treatment 1 and 53 µL for treatment solution 2.
- % of co-solvent per sample (mL/g x 100 %): Reported as < 0.1 % (actual 0.01%)
- Application method: Added drop wise to the water layer (equipment not reported)

EXPERIMENTAL WATER CONDITIONS
- Temperature: 20 ± 2 °C
- pH river water: 8.2 at study initiation and 8.3 at study termination.
- pH pond water: 8.4 at study initiation and 8.3 at study termination.
- Redox potential river water (mV): 374 at study initiation and 259 at study termination.
- Redox potential pond water (mV): 378 at study initiation and 278 at study termination.
- Oxygen concentration river water (mg O2/L): 6.8 at study initiation and termination.
- Oxygen concentration pond water (mg O2/L): 6.7 at study initiation and termination.

EXPERIMENTAL SEDIMENT CONDITIONS
- Redox potential river sediment (mV): -497 at study initiation and -402 at study termination
- Redox potential pond sediment (mV): -425 at study initiation and -389 at study termination
- Biomass river sediment (mg carbon/100 g soil): 77.84 at study initiation and 41.73 at study termination
- Biomass pond sediment (mg carbon/100 g soil): 119.16 at study initiation and 44.84 at study termination

CALCULATIONS
The rate of dissipation of the test substance from the water phase and the rate of degradation in the total aquatic sediment system were calculated assuming first order (one and two compartment) kinetics.
Reference substance:
not required
Test performance:
The overall mass balance for the radiolabelled test substance in the river (Rhine) and pond (Rheinfelden) test systems were 95.0 % and 97.1 % AR, respectively, during the 100-day study, which satisfied the guideline requirement of maintaining material balance between 90 % and 110 % AR.
Compartment:
natural water / sediment: freshwater
% Recovery:
95.99
St. dev.:
5.28
Remarks on result:
other: mean recovery of applied radioactivity (RA) to the 'river' water/sediment system
Remarks:
Total recovery is the sum of activity in the surface water, sediment extracts, sediment residue following combustion and that trapped as 14CO2 in the 2N NaOH traps
Compartment:
natural water / sediment: freshwater
% Recovery:
97.14
St. dev.:
7.55
Remarks on result:
other: mean recovery of applied radioactivity (RA) to the 'pond' water/sediment system
Remarks:
Total recovery is the sum of activity in the surface water, sediment extracts, sediment residue following combustion and that trapped as 14CO2 in the 2N NaOH traps
% Degr.:
ca. 75.93
Parameter:
radiochem. meas.
Sampling time:
100 d
Remarks on result:
other: degradation as total loss of parent substance in the river system (Rhine).
Remarks:
Standard deviation not reported. A detailed overview of the degradation is presented in 'Any other information on results incl. tables'.
% Degr.:
ca. 84.06
Parameter:
radiochem. meas.
Sampling time:
100 d
Remarks on result:
other: degradation as total loss of parent substance in the pond system (Rheinfelden) (93.84% in water and 90.22% in sediment).
Remarks:
Standard deviation not reported. A detailed overview of the degradation is presented in 'Any other information on results incl. tables'.
Compartment:
natural water / sediment: freshwater
DT50:
36.3 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: in Rhine river total water-sediment system
Compartment:
natural water / sediment: freshwater
DT50:
25.7 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: in Rheinfelden pond total water-sediment system
Compartment:
natural water: freshwater
DT50:
11 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: in Rhine river water
Compartment:
natural water: freshwater
DT50:
8.4 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: in Rheinfelden pond water
Compartment:
natural sediment: freshwater
DT50:
12.6 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: in Rhine river sediment
Compartment:
natural sediment: freshwater
DT50:
15.6 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: in Rheinfelden pond sediment
Transformation products:
yes
Details on transformation products:
- Formation and decline of each transformation product during test: Dissipation of the parent molecule from the water phase proceeded mainly via adsorption to the sediment matrix where the compound was reduced to the denitro metabolite M3. This transformation product could be extracted mainly by a harsh extraction method using acetonitrile-water and acetonitrile-hydrochloric acid under reflux. It reached maximum concentrations of 37.0 % AR at day 58 and decreased to 35.4 % AR at day 100 in the river and 47.4% AR at day 42 and 45.5% AR at day 100 in the pond sediment, respectively. The other metabolite identified as M1 was observed in both water and sediment of the river system at levels below 5 % AR beginning at day 42. In the pond system it was present beginning at day 8 at lower levels.
Evaporation of parent compound:
no
Volatile metabolites:
no
Residues:
not specified
Details on results:
The results are tabulated in 'Any other information on results incl. tables'.

TEST CONDITIONS
- Indication of test material adsorbing to walls of test apparatus: No

DEGRADATION OF THE TEST SUBSTANCE
In both systems the radioactivity in the water phase decreased over the incubation period reaching 18.4 (river) and 6.2 (pond) of the applied radioactivity at day 100. In the sediment the amount of non-extractable radioactivity increased continuously to a level of 13.8 % AR (river) and 15.0 % AR (pond). All volatile radioactivity was characterized as carbon dioxide which reached 6.3 % AR (river) and 9.3 % AR (pond) at day 100 and indicated some mineralisation. The distribution and nature of radioactivity for the incubation period in the river and pond water-sediment systems are presented in 'Any other information on results incl. tables'.

WATER RESIDUES
Water residues declined with time in all test systems.
- Total water residues at 0 days after treatment: River (Rhine): 102.01%; Pond (Rheinfelden): 104.69%
- Total water residues at end of study: River (Rhine): 18.39%; Pond (Rheinfelden): 6.16%

EXTRACTABLES RESIDUES
Extractable residues decreased with time in all test systems.
- Total extractable residues at 0 days after treatment (including radioactivity in water phase): River (Rhine): 103.47%; Pond (Rheinfelden): 107.10%
- Total extractable residues at end of study (including radioactivity in water phase): River (Rhine): 70.75%; Pond (Rheinfelden): 64.77%

BOUND RESIDUES
Bound residues increased throughout the incubation period.
- Bound residues at end of study: River (Rhine): 13.79%; Pond (Rheinfelden): 14.98%

VOLATILISATION
14-CO2 was evolved as volatile products throughout the course of the study
- 14-CO2 evolved at end of study: River (Rhine): 6.34%; Pond (Rheinfelden): 9.28%
- Other volatiles: No other volatile radioactivity detected

NON-EXTRACTABLE RESIDUES
The un-extracted residue remaining after the harsh extraction was characterized by organic matter fractionation, i.e. fulvic acid, humic acid, and humin. Most of the radioactivity remaining after the harsh extraction was characterized as being associated with the fulvic acid fraction, i.e. a maximum of 4.38 and 8.64 % AR for the river and pond aquatic systems, respectively.

DISTRIBUTION COEFFICIENTS
Based on the relative distribution of the test substance to water/sediment for Rhine river and for pond aquatic systems, distribution coefficients kd for any time can be estimated taking the dimensions of the systems into account. After decantation/centrifugation about 540 mL water phase represented the water layer. The mean dry weight of the sediments was 122 g per flask for the river and for the pond system corresponding to about 200 g wet sediments taken for extraction after decanting. 52 µg radiolabelled test substance was applied per flask. After 16 days of incubation equilibrium was reached in both systems and about 34% of the applied radioactivity were found in both systems in the water phase as well as in the sediment. The concentrations of the radiolabelled test substance in the sediments can be calculated to be about 89 µg/kg and the concentration of radiolabelled test substance in water can be calculated to about 33 µg/litre. Thus, a mean sediment/water distribution coefficient can be estimated to be kd = 2.7 mL/g.

Table: Radioactivity distribution of the radiolabelled test substance and metabolites in river (Rhine) aquatic system as percent of applied dose (values as percentage of applied radioactivity)

Fraction

Incubation Time (days), values are given in % of the radioactivity applied(1)

0

1

3

8

16

42

58

80

100

Water

102.01

81.46

78.08

63.62

37.22

21.9

22.9

16.2

11.8

Extractables before Harsh Extraction(2)

1.46

13.49

23.44

28.41

33.57

33.69

27.25

26.70

26.24

Harsh Extractions

n.d.

n.d.

n.d.

n.d.

18.80

25.25

32.89

26.52

26.12

Total-Extractables

1.46

13.49

23.44

28.41

52.37

58.94

60.14

53.22

52.36

Non-Extractables after Harsh Extractions

0.28

0.35

2.01

5.29

5.05

7.53

5.79

12.59

13.79

Volatiles

n.d.

0.02

0.09

0.88

1.40

1.07

1.56

2.41

6.34

Recovery(3)

103.75

95.32

103.62

98.20

96.04

94.19

93.98

87.90

90.88

Average Recovery ± SD

95.99 ± 5.28

n.d. - not determined; SD - standard deviation

1) duplicate samples performed for each water/sediment system for each sampling interval

2) including Soxhlet extractions

3) recovery is determined by LSC analysis of the individual fractions; Recovery = % water fraction + % extractables + % non-extractables + % volatiles

 

Table: Radioactivity distribution of the radiolabelled test substance and metabolites in pond (Rheinfelden) aquatic system as percent of applied dose (values as percentage of applied radioactivity)

Fraction

Incubation Time (days), values are given in % of the radioactivity applied(1)

0

1

3

8

16

42

58

80

100

Water

104.69

85.21

75.85

53.9

33.6

12.3

26.7

2.97

6.16

Extractables before Harsh Extraction(2)

2.41

14.92

21.15

25.58

31.16

25.65

23.96

17.89

23.90

Harsh Extractions

n.d.

n.d.

n.d.

12.91

16.97

39.64

25.93

39.36

34.71

Total-Extractables

2.41

14.92

21.15

38.49

48.13

65.29

49.89

57.25

58.61

Non-Extractables after Harsh Extractions

0.73

0.98

3.53

4.36

7.50

7.08

21.86

25.32

14.98

Volatiles

n.d.

0.02

0.07

0.05

0.44

2.19

5.70

3.17

9.28

Recovery(3)

107.83

101.13

100.60

99.92

92.21

88.96

105.95

88.67

89.03

Average Recovery ± SD

97.14 ± 7.55

n.d. - not determined; SD - standard deviation

1) duplicate samples performed for each water/sediment system for each sampling interval

2) including Soxhlet extractions

3) recovery is determined by LSC analysis of the individual fractions; recovery = % water fraction + % extractables + % non-extractables + % volatiles

 

Table: Distribution of the radiolabelled test substance and metabolites in the water fraction of the river (Rhine) system (values as percentage of applied radioactivity)

Test substance

Incubation Time (days), values are given in % of the radioactivity applied

0

1

3

8

16

42

58

80

100

Parent compound

102.01

81.46

78.08

63.62

37.22

21.92

22.87

16.16

11.83

M1

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

3.01

1.47

3.52

3.96

M3

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.=lower than detection limit

 

Table: Distribution of the radiolabelled test substance and metabolites in the sediment fraction of the river (Rhine) system (values as percentage of applied radioactivity)

Test substance

Incubation Time (days), values are given in % of the radioactivity applied

0

1

3

8

16

42

58

80

100

Parent compound

1.46

13.49

23.44

27.21

33.57

21.86

13.63

12.66

12.24

M1

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

4.09

3.97

4.69

M3

<l.d.

<l.d.

<l.d.

<l.d.

18.80

33.00

36.98

30.67

35.44

<l.d.=lower than detection limit

 

Table: Distribution of the radiolabelled test substance and metabolites in the total water/sediment fraction of the river (Rhine) system (values as percentage of applied radioactivity) 

Test substance

Incubation Time (days), values are given in % of the radioactivity applied(1)

0

1

3

8

16

42

58

80

100

Parent compound

103.47

94.95

101.52

90.83

70.79

43.78

36.50

28.82

24.07

M1

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

3.01

5.56

7.49

8.65

M3

<l.d.

<l.d.

<l.d.

<l.d.

18.80

33.00

36.98

30.67

35.44

<l.d.= lower than detection limit

1) total water/sediment fraction values are sum of water fraction and sediment fraction values

 

Table: Distribution of the radiolabelled test substance and metabolites in the water fraction of the pond (Rheinfelden) system (values as percentage of applied radioactivity)

Test substance

Incubation Time (days), values are given in % of the radioactivity applied

0

1

3

8

16

42

58

80

100

Parent compound

104.69

85.21

75.85

53.66

33.56

12.32

26.66

2.93

6.16

M1

<l.d.

<l.d.

<l.d.

2.23

1.05

2.08

1.27

<l.d.

<l.d.

M3

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

Not specified metabolite

<l.d.

<l.d.

<l.d.

1.13

1.53

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.= lower than detection limit

 

Table: Distribution of the radiolabelled test substance and metabolites in the sediment fraction of the pond (Rheinfelden) system (values as percentage of applied radioactivity)

Test substance

Incubation Time (days), values are given in % of the radioactivity applied

0

1

3

8

16

42

58

80

100

Parent compound

2.41

14.92

21.15

22.28

31.16

14.51

13.87

9.75

9.78

M1

<l.d.

<l.d.

<l.d.

1.15

<l.d.

3.39

2.45

2.55

3.39

M3

<l.d.

<l.d.

<l.d.

15.06

16.22

47.39

32.04

46.28

45.45

Not specified metabolite

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

<l.d.

0.48

<l.d.

<l.d.

<l.d.= lower than detection limit

 

Table: Distribution of the radiolabelled test substance and metabolites in the total water/sediment fraction of the pond (Rheinfelden) system (values as percentage of applied radioactivity)

Test substance

Incubation Time (days), values are given in % of the radioactivity applied(1)

0

1

3

8

16

42

58

80

100

Parent compound

107.10

100.13

97.00

75.94

64.72

26.83

47.70

15.25

15.94

M1

<l.d.

<l.d.

<l.d.

3.38

1.05

5.47

3.72

2.55

3.39

M3

<l.d.

<l.d.

<l.d.

27.97

32.44

87.03

55.76

84.42

80.16

Not specified metabolite

<l.d.

<l.d.

<l.d.

1.13

1.53

<l.d.

0.48

<l.d.

<l.d.

<l.d.= lower than detection limit

1) total water/sediment fraction values are sum of water fraction and sediment fraction values

 

Table: Summary of recovery of radioactivity by harsh extraction and organic matter fractionation (values as percentage of applied radioactivity)

Incubation Days

Harsh extraction

Organic matter fractionation

ACN/H20

ACN/HCL

Fulvic acids

Humic Acids

Insoluble (humin)

River (Rhine) aquatic system

1

n.p.

n.p.

n.p.

n.p.

n.p.

3

n.p.

n.p.

n.p.

n.p.

n.p.

8

n.p.

n.p.

n.p.

n.p.

n.p.

16

11.80

7.00

1.34

0.37

0.31

42

15.65

9.60

1.99

0.29

0.67

58

17.64

15.25

2.79

0.93

0.00

80

14.64

11.88

4.09

0.45

0.37

100

7.82

18.30

4.38

0.51

0.15

Pond (Rheinfelden) aquatic system

1

n.p.

n.p.

n.p.

n.p.

n.p.

3

n.p.

n.p.

n.p.

n.p.

n.p.

8

6.51

6.40

1.29

0.44

0.15

16

9.80

7.17

2.36

0.48

0.05

42

17.65

21.99

n.p.

n.p.

n.p.

58

8.00

17.93

7.82

1.08

0.83

80

18.64

20.72

8.64

0.45

0.64

100

17.01

17.70

6.72

0.05

0.74

n.p. Not performed

Table: Half-lives of the radiolabelled test substance in aquatic systems

 

Total System

Water

Sediment

(days)

DT50

DT90

DT50

DT90

DT50

DT90

River

36.3

317.4

11.0

132.3

12.6

41.9

Pond

25.7

130.9

8.4

76.9

15.6

51.9

Validity criteria fulfilled:
yes
Conclusions:
The transformation of radiolabelled test substance in aerobic aquatic sediments was assessed in two test systems (water pH characteristics pH 7.9 - 8.3) that were continuously aerated in the dark at a temperature of 20 ± 2°C for 100 days. The overall mass balance for the radiolabelled test substance in the river (Rhine) and pond (Rheinfelden) test systems were 95.0 % and 97.1 % AR, respectively, during the 100-day study, which satisfied the guideline requirement of maintaining material balance between 90 % and 110 % AR. The test substance was shown to partition from the water phase to the sediment with dissipation half-lives ranging from 8.4 to 11.0 days. Transformation of the test substance in the water phase and sediment resulted in degradation half-lives ranging from 25.7 to 36.3 days and the formation of three products, including CO2. Three transformation products were identified as M3 and M1 by co-chromatography using an HPLC and TLC system. Further degradation resulted in formation of polar products (in bound residues), mineralization and strong binding to fractions of the sediment organic matter.
Executive summary:

The biodegradation of the test item in water/sediments systems was investigated in a study according to SETAC 8.2 and BBA part IV, 5-1, and in compliance with GLP criteria. In this study, the rate and route of degradation of the test substance radiolabelled in the thiazole ring, was investigated in two aquatic sediments and associated overlaying water collected from a river (Rhineat Mohlin) and a pond (Judenweiher at Rheinfelden) locations. Water was treated with the radiolabelled test substance to achieve target concentrations of 0.1 mg/L in the water phase (based on the maximum single label application rate of 300 g a.s./ha). The systems were incubated under aerobic conditions in the laboratory and maintained in dark conditions at 20°C for up to 100 days. Duplicate samples were taken for analysis at nine time intervals (0, 1, 3, 8, 16, 42, 58, 80 and 100 days after treatment). Average mass balance for the river (Rhine) aquatic system ranged from 87.90 to 103.75 % of the applied radioactivity (AR). Similarly, average mass balance for the pond (Rheinfelden) aquatic system ranged from 88.96 to 107.83 % AR over the course of the 100-day study. Half-life (DT50) for the dissipation of the radiolabelled test substance in the water phase ranged from 8.4 to 11.0 days for the aquatic systems. The half-life of the degradation of the radiolabelled test substance in the total aquatic sediment systems ranged from 25.7 to 36.3 days for aquatic systems. The DT50and DT90 values were determined by applying first order one and two compartment reaction kinetics. The 14-CO2 evolution due to mineralization of the radiolabelled test substance ranged from 0.02 to 6.34 % AR and 0.02 to 9.28 % AR, for the river and pond aquatic systems, respectively. No other volatile components were reported. Transformation products of radiolabelled test substance was observed in the aquatic aerobic sediment systems as evidenced by HPLC profiling. Metabolites M1 and M3 were identified by HPLC and TLC co-chromatography with authentic reference standards. Average non-extractable residues increased to levels of 13.8 % and 15.0 % AR at 100 DAT in the river and pond aquatic systems, respectively. Radioactivity remaining in the PES samples after soft and harsh extractions, was characterized by fractionation into humin, humic acid and fulvic acid fractions in which the majority of the bound radioactivity was associated with the fulvic acid fraction, i.e. a maximum of 4.38 and 8.64 % AR for the river and pond aquatic systems, respectively.

Endpoint:
biodegradation in water and sediment: simulation testing, other
Remarks:
natural water/sediment systems (pond and river).
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 308 (Aerobic and Anaerobic Transformation in Aquatic Sediment Systems)
Version / remarks:
April 2002
Qualifier:
according to
Guideline:
EPA OPPTS 835.4300 (Aerobic Aquatic Metabolism)
Version / remarks:
October 2008
GLP compliance:
yes (incl. certificate)
Radiolabelling:
yes
Remarks:
14C-labelled at position 2 of thiazole ring or position 4 of the oxadiazine ring
Oxygen conditions:
aerobic
Inoculum or test system:
natural water / sediment: freshwater
Details on source and properties of surface water:
SOURCE AND SAMPLING OF RIVER WATER
- Source: Taunton River at Bridgewater (Massachusetts, USA)
- Site description: River - sandy loam sediment
- Geographical region / Global co-ordinates: North Eastern U.S.A / 41º56.863N - 70º56.201W.
- Date of collection: 24 October 2013
- Sampling: Water was sampled down to a depth of 30 cm above the sediment with a container. Water samples were clear.
- Storage conditions after sampling: Stored refrigerated in open container with access to air.

RIVER WATER PROPERTIES
- Oxygen concentration: 7.5 % at sampling
- pH: 6.0 at sampling
- Redox potential: 152.1 mV at the start of acclimation
Details on source and properties of sediment:
SOURCE AND SAMPLING OF SEDIMENT
- Source: Same as water
- Site description: Same as water
- Geographical region / Global co-ordinates: Same as water
- Date of collection: Same as water
- Sampling: 0 - 7.6 cm of sediment was sampled using a shovel and sieved to 2 mm.
- Storage conditions after sampling: Stored refrigerated in separate open container with access to air

PROPERTIES OF RIVER SEDIMENT
- % Clay (<2 µm): 5
- % Silt (50-2 µm): 49
- % Sand (2000-50 µm: 46
- Soil type (USDA): Sandy loam
- pH (water) in 1:1 sediment:water: 5.5
- Redox potential: 155.7 mV at the start of acclimation
- Organic matter (%): 6.6
- Organic carbon (%): 3.8
- Cation Exchange Capacity (meq/100g soil): 7.1
- Nitrogen (total, %): 0.23
- Phosphorus (total, mg/kg): 78
- Moisture (w/w %): 60.34
Duration of test (contact time):
100 d
Initial conc.:
0.1 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
CO2 evolution
radiochem. meas.
Details on study design:
TEST SYSTEM
- Apparatus: Glass flask, moist air bubbled through system, connections made with glass and Teflon® tubing
- Number of replicates: 2 replicates per radiolabel for each sampling interval
- Sediment condition: Fresh, passed through 2 mm sieve prior to use
- Water condition: Fresh, collected before sediment, allowed any particulates to settle before use.
- Control conditions: Not applicable
- Incubation conditions: Aerobic - aerated continuously with ambient hydrated air.

SAMPLE SIZES
- River water: 212 mL (resulting in 4:1 water: sediment ratio)
- Pond water: 109 g wet weight (52 g dry weight)

TEST MATERIAL APPLICATION
- Solvent: Acetonitrile
- Volume of test solution used per treatment: 230 µL for [oxadiazine-4-14C]-test substance and 235 µL for [thiazole-2-14C]-test substance
- % of co-solvent per sample (mL/g x 100 %): < 0.1 %
- Application method: Hamilton syringe followed by gentle mixing

EXPERIMENTAL WATER CONDITIONS
- Temperature: 20 ± 2 °C
- Light: Continuous darkness
- pH river water: 6.5 at acclimation and study initiation and 4.4 at study termination.
- Redox potential river water (mV): 152.1 at acclimation, 92.5 at study initiation and 238.4 at study termination.
- Oxygen concentration river water (%): 5.3 at acclimation and study initiation and 4.0 at study termination

EXPERIMENTAL SEDIMENT CONDITIONS
- Redox potential river sediment (mV): 155.7 at acclimation, -78.0 at study initiation and 19.3 at study termination.
- Biomass river sediment (mg carbon/100 g soil): 13.7 at study initiation and 11.2 at study termination

CALCULATIONS
The rate of dissipation of the test substance from the water phase and the rate of degradation in the total aquatic sediment system were calculated using the Simple First-Order (SFO) kinetic model.

STATISTICS
The dissipation rate (DT50) from water and the degradation rate (DegT50) of the parent in the total system were determined from the best-fit model (SFO, CAKE, Version 1.4) based on acceptable visual fits (high correlation coefficient, r2) as well as statistical evaluation (lowest X2 % error).
Reference substance:
not required
Test performance:
The overall mass balance for the [oxadiazine-4-14C]-test substance and [thiazole-2-14C]-test substance in the Taunton River test system was 95.8 % and 97.9 % AR, during the 100-day study, which satisfied the guideline requirement of maintaining material balance between 90 % and 110 % AR.
Compartment:
other:
% Recovery:
97.93
St. dev.:
4.18
Remarks on result:
other: Recovery rates ranged from 92.02 to 105.25% AR for river aquatic system
Remarks:
mean overall recovery (all samples) of applied radioactivity (RA) of [oxadiazine-4-14C]-test substance. Total recovery is the sum of activity in the surface water, sediment extracts, sediment residue following combustion and that trapped as 14CO2 in the 1M KOH traps.
Compartment:
natural water / sediment: freshwater
% Recovery:
95.81
St. dev.:
2.64
Remarks on result:
other: Recovery rates ranged from 91.26 to 103.06% AR for river aquatic system.
Remarks:
mean overall recovery (all samples) of applied radioactivity (RA) of [thiazole-2-14C]-test substance. Total recovery is the sum of activity in the surface water, sediment extracts, sediment residue following combustion and that trapped as 14CO2 in the 1M KOH traps.
% Degr.:
82.8
Parameter:
radiochem. meas.
Sampling time:
100 d
Remarks on result:
other: [oxadiazine-4-14C]-test substance parent compound degradation in the total water/sediment test system
% Degr.:
86.6
Parameter:
radiochem. meas.
Sampling time:
100 d
Remarks on result:
other: [thiazole-2-14C]-test substance parent compound degradation in the total water/sediment test system
Compartment:
natural water / sediment: freshwater
DT50:
29.2 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: [Oxadiazine-4-14C]-test substance rate of degradation in whole system
Compartment:
natural water / sediment: freshwater
DT50:
25.2 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: [Thiazole-2-14C]-test substance rate of degradation in whole system
Compartment:
natural water: freshwater
DT50:
19.2 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: [Oxadiazine-4-14C]-test substance rate of dissipation from the water phase.
Compartment:
natural water: freshwater
DT50:
14.8 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Remarks on result:
other: [Thiazole-2-14C]-test substance rate of dissipation from the water phase.
Transformation products:
yes
Details on transformation products:
- Formation and decline of each transformation product during test: In both water-sediment systems, during the aerobic phase, the major degradation products were M1, M2 and M3. Mineralization was a major route of aerobic degradation. With the oxadiazine ring or the thiazole ring labels, M3 reached maximum mean levels of 17.8 % and 17.0 % AR, at 70 days after treatment, and decreased to 16.2 % and 13.8 % AR, respectively, by the end of the study. Levels of M2 in the oxadiazine ring or the thiazole ring reached maximum mean levels of 3.54 % and 6.84 % AR at 48 days after treatment and decreased to 1.28 % and 0.67 % AR, respectively, by the end of the study. With the oxadiazine ring or the thiazole ring labels, M1 reached maximum mean levels of 5.98 % and 6.01 % AR, at 48 days after treatment, and decreased to 1.3 % and 2.27 % AR, respectively, by the end of the study. A fourth transformation product eluted at a retention time of approximately 2 to 3 min as a polar product, which did not correspond to any reference standard. This polar product was present at average maximum levels of 3.96 % at 48 days after treatment and 7.43 % AR at 14 days after treatment in the system treated with the oxadiazine ring or the thiazole ring (>5 % at one interval and in one label only), but decreased to 1.44 % and 0.18 % AR, respectively by the end of the study. The individual peaks of the polar product represented <5 % AR and were not considered further.
Evaporation of parent compound:
no
Volatile metabolites:
no
Remarks:
Negligible reaching a maximum of 0.58 % AR at 30 days after treatment
Residues:
not specified
Details on results:
The results are tabulated in 'Any other information on results incl. tables'.

TEST CONDITIONS
- Indication of test material adsorbing to walls of test apparatus: No

DEGRADATION OF THE TEST SUBSTANCE
Levels of the parent compound in the total water/sediment test system decreased from 96.6 and 94.5% AR at 0 days after treatment to 17.2 and 13.4 % AR at 100 days after treatment in the aerobic test system dosed with [oxadiazine-4-14C]-test substance and [thiazole-2-14C]-test substance, respectively.

WATER RESIDUES
Water residues declined with time in all test systems.
- Total water residues at 0 days after treatment: Oxadiazine label: Mean 94.90%; Thiazole label: Mean 93.7%
- Total water residues at end of study: Oxadiazine label: Mean 16.82%; Thiazole label: Mean 9.45%

EXTRACTABLES RESIDUES
Extractable residues decreased with time in all test systems.
- Total extractable residues at 0 days after treatment (including radioactivity in water phase): Oxadiazine label: Mean 96.56%; Thiazole label: Mean 94.51%
- Total extractable residues at end of study (including radioactivity in water phase): Oxadiazine label: Mean 41.84 %; Thiazole label: Mean 31.50 %

BOUND RESIDUES
The average amount of bound residue increased from 0.51% and 0.24% AR at 0 days after treatment to 48.8% and 51.32% AR by 100 days after treatment in the test systems dosed with [oxadiazine-4 14C]-test substance and [thiazole-2-14C]-test substance, respectively. The un-extracted residue remaining after the harsh extraction (of the 100 days after treatment sample) was characterized by organic matter fractionation, i.e. fulvic acid, humic acid, and humin. The most of the radioactivity remaining after the harsh extraction was characterized as being bound to the humin fraction, i.e., from 18.2 to 19.5% AR.

VOLATILISATION
Significant amount of 14CO2 was evolved as volatile products throughout the course of the study.
- 14-CO2 evolved at end of study: Oxadiazine label: Mean 6.42 %; Thiazole label: Mean 11.96 %
- Other volatiles: Negligible reaching a maximum of 0.58% AR at 30 days after treatment

NON-EXTRACTABLE RESIDUES
The un-extracted residue remaining after the harsh extraction was characterized by organic matter fractionation, i.e. fulvic acid, humic acid, and humin. Most of the radioactivity remaining after the harsh extraction was characterized as being associated with the fulvic acid fraction, i.e. a maximum of 4.38 and 8.64 % AR for the river and pond aquatic systems, respectively.

Table: Mass Balance and Distribution of Radioactivity in the Taunton River Sediment System Tested with [oxadiazine-4 14C]-test substance (values as percentage of Applied Radioactivity)

Fraction

Replicate

Incubation Time (DAT)

0

7

14

30

48

70

100

Surface Water (%)

A

96.00

69.14

65.46

33.32

26.62

11.83

16.69

B

93.80

65.12

60.32

34.47

29.39

18.45

16.96

Mean

94.90

67.13

62.89

33.89

28.00

15.14

16.82

Sediment Extraction (%)

A

1.28

20.38

25.74

23.78

26.40

26.38

25.06

B

2.04

21.77

27.03

25.36

28.00

30.11

24.96

Mean

1.66

21.1

26.4

24.6

27.2

28.3

25.0

Total Extractable

Mean

96.56

88.21

89.28

58.46

55.20

43.38

41.84

Non-Extractable (%)

A

0.40

5.56

9.69

33.52

42.37

59.09

46.48

B

0.62

5.03

12.35

29.89

37.07

49.47

51.06

Mean

0.51

5.29

11.02

31.70

39.72

54.28

48.77

14CO2(%)

A

NA

0.09

0.22

2.04

4.71

7.19

6.42

B

NA

0.09

0.22

2.04

4.71

7.19

6.42

Mean

NA

0.09

0.22

2.04

4.71

7.19

6.42

Other Volatiles (%)

A

NA

ND

ND

0.58

ND

0.03

ND

B

NA

ND

ND

0.58

ND

0.03

ND

Mean

NA

ND

ND

0.58

ND

0.03

ND

Mass Balance (%)

A

97.68

95.18

101.12

93.24

100.10

104.52

94.65

B

96.47

92.02

99.92

92.34

99.16

105.25

99.40

Mean

97.07

93.60

100.52

92.79

99.63

104.89

97.02

Mean ± SD

97.93 ± 4.18

NA - not applicable; ND - not detected or < 0.01 % AR; VOC - volatile organic compounds; SD – standard deviation

All reported values are rounded. All calculations are based on unrounded values.

Mass balance is determined by LSC analysis of the individual fractions; Mass balance = % water fraction + % extractable +% bound +% 14CO2 + % VOC

Total radioactivity applied to the water/sediment test systems at day 0 = 6,864,671 dpm; Specific Activity [oxadiazine-4-14C]-test substance used in the water/sediment study = 311,022 dpm/µg.

Total µg applied to the water/sediment test systems = 21.2 µg; Total volume of water in the test systems = 212 mL. Nominal test system ppm = 21.2 µg/212 mL = 0.10 µg/mL = 0.10 ppm

Table: Mass Balance and Distribution of Radioactivity in the Taunton River Sediment System Tested with [thiazole-2-14C]-test substance (values as percentage of Applied Radioactivity)

Fraction

Replicate

Incubation Time (DAT)

0

7

14

30

48

70

100

Surface Water (%)

A

92.84

64.22

56.11

27.91

25.54

18.56

10.35

B

94.58

64.89

56.01

27.39

21.00

19.67

8.56

Mean

93.71

64.56

56.06

27.65

23.27

19.11

9.45

Sediment Extraction (%)

A

0.93

23.22

27.93

22.38

27.52

30.59

21.90

B

0.68

20.65

27.03

23.33

24.77

31.10

22.19

Mean

0.80

21.94

27.48

22.85

26.15

30.85

22.04

Total Extractable

Mean

94.51

86.50

83.54

50.50

49.42

49.96

31.50

Non-Extractable (%)

A

0.25

7.82

14.52

33.89

33.90

43.61

47.88

B

0.22

5.55

14.46

33.14

37.27

33.20

54.75

Mean

0.24

6.68

14.49

33.51

35.59

38.41

51.32

14CO2 (%)

A

NA

0.17

1.23

9.04

10.90

10.30

11.96

B

NA

0.17

1.23

9.04

10.90

10.30

11.96

Mean

NA

0.17

1.23

9.04

10.90

10.30

11.96

Other Volatiles (%)

A

NA

ND

0.02

0.05

0.03

ND

ND

B

NA

ND

0.02

0.05

0.03

ND

ND

Mean

NA

ND

0.02

0.05

0.03

ND

ND

Mass Balance (%)

A

94.02

95.43

99.82

93.27

97.90

103.06

92.09

B

95.48

91.26

100.33

92.95

93.98

94.27

97.46

Mean

94.75

93.35

100.07

93.11

95.94

98.67

94.78

Mean ± SD

95.81 ± 2.64

NA - not applicable, ND - not detected or < 0.01 % AR, VOC - volatile organic compounds; SD – standard deviation

All reported values are rounded. All calculations are based on unrounded values.

Mass balance is determined by LSC analysis of the individual fractions. Mass balance = % water fraction + % extractable +% bound +% 14CO2 + % VOC.

Total radioactivity applied to the water/sediment test systems at day 0 = 6,864,671 dpm

Specific Activity [oxadiazine-4-14C]-test substance used in the water/sediment study = 311,022 dpm/µg

Total µg applied to the water/sediment test systems = 21.2 µg; Total volume of water in the test systems = 212 mL; Nominal test system ppm = 21.2 µg/212 mL = 0.10 µg/mL = 0.10 ppm

 

Table: Distribution of [oxadiazine-4-14C]-test substance and Metabolites in the Water Fraction of the Taunton River Aerobic Test Systems (values as percentage of Applied Radioactivity)

[oxadiazine-4-14C]-test substance

Replicate

Sampling time (DAT)

0

7

14

30

48

70

100

Parent compound

A

96.00

64.56

57.70

26.32

18.56

6.78

11.18

B

93.80

62.29

53.47

30.86

16.72

11.87

9.95

Mean

94.90

63.42

55.59

28.59

17.64

9.32

10.57

Polar(a)

A

ND

1.98

2.68

0.72

1.58

0.99

0.81

B

ND

1.05

2.45

ND

3.68

0.72

1.56

Mean

ND

1.52

2.57

0.36

2.63

0.86

1.18

Unspecified metabolite

A

ND

ND

ND

ND

ND

ND

ND

B

ND

ND

ND

ND

ND

ND

ND

Mean

ND

ND

ND

ND

ND

ND

ND

M3

A

ND

ND

ND

0.55

1.34

1.70

1.10

B

ND

ND

ND

ND

1.39

0.96

1.86

Mean

ND

ND

ND

0.27

1.37

1.33

1.44

M2

A

ND

ND

2.16

1.95

2.13

0.99

0.98

B

ND

ND

1.15

1.88

3.43

1.63

1.14

Mean

ND

ND

1.65

1.91

2.78

1.31

1.06

M1

A

ND

2.61

2.91

2.22

3.00

1.36

1.57

B

ND

1.78

2.34

1.74

4.17

1.34

1.38

Mean

ND

2.20

2.62

1.98

3.59

1.35

1.47

Others(b)

A

ND

ND

ND

1.57

ND

ND

1.04

B

ND

ND

0.90

ND

ND

1.92

1.08

Mean

ND

ND

0.46

0.78

ND

0.96

1.06

a) HPLC and/or TLC chromatograms demonstrate that freely extractable “polars” were multi component and each were <5 % applied radioactivity (AR).

b) “Others” consists of multiple components and includes all peaks not quantified in the table above; HPLC and/or TLC chromatograms demonstrate that multi component “others” were < 5 % AR.

ND - Not Detected or < 0.1 %AR; % metabolite water = (% by HPLC in water) × (% of applied radioactivity in the water fraction)

Note: All reported values are rounded. All calculations are based on unrounded values.

 

Table: Distribution of [oxadiazine-4-14C]-test substance and Metabolites in the Sediment Extractable Fraction of the Taunton River Aerobic Test System (values as percentage of Applied Radioactivity)

[oxadiazine-4-14C]-test substance

Replicate

Sampling time (DAT)

0

7

14

30

48

70

100

Parent compound

A

1.28

17.81

15.30

13.78

11.20

9.43

6.66

B

2.04

19.45

20.35

12.75

12.10

7.45

6.44

Mean

1.66

18.63

17.83

13.26

11.65

8.44

6.55

Polar(a)

A

ND

ND

1.55

ND

1.60

ND

0.51

B

ND

ND

0.72

0.69

1.07

ND

ND

Mean

ND

ND

1.1

0.3

1.3

ND

0.3

Unspecified metabolite

A

ND

ND

ND

ND

ND

ND

ND

B

ND

ND

ND

ND

ND

ND

ND

Mean

ND

ND

ND

ND

ND

ND

ND

M3

A

ND

2.23

3.07

7.77

10.78

12.05

14.23

B

ND

2.32

5.10

10.42

11.34

20.82

15.13

Mean

ND

2.27

4.08

9.09

11.06

16.44

14.68

M2

A

ND

ND

0.45

0.64

0.79

0.55

0.44

B

ND

ND

ND

ND

0.74

ND

ND

Mean

ND

ND

0.23

0.32

0.76

0.27

0.22

M1

A

ND

ND

1.63

1.60

2.04

2.44

1.47

B

ND

ND

0.87

1.49

2.75

1.84

1.35

Mean

ND

ND

1.33

1.55

2.39

2.14

1.41

Others(b)

A

ND

0.34

3.74

ND

ND

1.42

1.74

B

ND

ND

ND

ND

ND

ND

2.04

Mean

ND

0.17

1.87

ND

ND

0.71

1.89

a) HPLC and/or TLC chromatograms demonstrate that freely extractable “polars” were multi component and each were < 5 % applied radioactivity (AR).

b) “Others” consists of multiple components and includes all peaks not quantified in the table above; HPLC and/or TLC chromatograms demonstrate that multi component “others” were < 5% AR.

ND - Not Detected or < 0.1%AR; % metabolite water = (% by HPLC in water) × (% of applied radioactivity in the water fraction)

Note: All reported values are rounded. All calculations are based on unrounded values.

  

Table: Distribution of [thiazole-2-14C]-test substance and Metabolites in the Water Fraction of the Taunton River Aerobic Test System (values as percentage of Applied Radioactivity)

[thiazole-2-14C]-test substance

Replicate

Sampling time (DAT)

0

7

14

30

48

70

100

Parent compound

A

92.84

62.22

47.25

15.96

14.32

13.13

6.74

B

94.58

60.49

47.32

21.52

12.00

13.61

6.87

Mean

93.71

61.35

47.29

18.74

13.16

13.37

6.80

Polar(a)

A

ND

1.13

4.11

4.39

ND

0.42

ND

B

ND

1.27

4.96

0.84

1.47

ND

ND

Mean

ND

1.20

4.54

2.61

0.74

0.21

ND

Unspecified metabolite

A

ND

ND

ND

ND

ND

ND

ND

B

ND

ND

ND

ND

ND

ND

ND

Mean

ND

ND

ND

ND

ND

ND

ND

M3

A

ND

ND

ND

1.01

ND

0.84

1.23

B

ND

ND

ND

0.68

ND

1.18

0.74

Mean

ND

ND

ND

0.85

ND

1.01

0.98

M2

A

ND

ND

1.14

2.08

6.77

1.94

0.72

B

ND

1.73

1.05

1.62

4.70

1.94

0.25

Mean

ND

0.87

1.10

1.85

5.74

1.94

0.49

M1

A

ND

0.89

1.46

2.50

4.45

2.23

1.15

B

ND

1.40

0.72

2.73

2.83

2.40

0.71

Mean

ND

1.14

1.09

2.61

3.64

2.31

0.93

Others(b)

A

ND

ND

2.15

1.97

ND

ND

0.50

B

ND

ND

1.95

ND

ND

0.54

ND

Mean

ND

ND

2.05

0.98

ND

0.27

0.25

a) HPLC and/or TLC chromatograms demonstrate that freely extractable “polars” were multi component and each were < 5 % applied radioactivity (AR).

b) “Others” consists of multiple components and includes all peaks not quantified in the table above; HPLC and/or TLC chromatograms demonstrate that multi component “others” were < 5 % AR.

ND - Not Detected or < 0.1 %AR; % metabolite water = (% by HPLC in water) × (% of applied radioactivity in the water fraction)

Note: All reported values are rounded. All calculations are based on unrounded values.

  

Table: Distribution of [thiazole-2-14C]-test substance in the Sediment Extractables Fraction of the Taunton River Aerobic Test System (values as percentage of Applied Radioactivity)

[thiazole-2-14C]-test substance

Replicate

Sampling time (DAT)

0

7

14

30

48

70

100

Parent compound

A

0.93

19.67

18.50

7.04

12.17

10.12

6.89

B

0.68

17.21

18.37

11.58

7.04

13.65

6.32

Mean

0.80

18.44

18.43

9.31

9.61

11.89

6.60

Polar(a)

A

ND

ND

3.55

1.08

1.49

ND

ND

B

ND

ND

2.17

0.74

1.04

ND

0.36

Mean

ND

ND

2.89

0.91

1.26

ND

0.18

Unspecified metabolite

A

ND

ND

ND

ND

ND

ND

ND

B

ND

ND

ND

ND

ND

ND

ND

Mean

ND

ND

ND

ND

ND

ND

ND

M3

A

ND

3.55

2.99

10.96

10.68

16.41

12.49

B

ND

2.67

2.48

9.26

12.92

15.64

13.13

Mean

ND

3.11

2.74

10.11

11.80

16.03

12.81

M2

A

ND

ND

ND

0.85

1.22

ND

0.38

B

ND

ND

ND

ND

0.97

ND

ND

Mean

ND

ND

ND

0.43

1.10

ND

0.19

M1

A

ND

ND

1.34

1.63

1.95

2.64

1.32

B

ND

ND

1.07

1.74

2.80

1.81

1.37

Mean

ND

ND

1.20

1.69

2.37

2.23

1.34

Others(b)

A

ND

ND

1.49

0.81

ND

1.42

0.83

B

ND

ND

4.53

ND

ND

ND

1.01

Mean

ND

ND

3.01

0.40

ND

0.71

0.92

a) HPLC and/or TLC chromatograms demonstrate that freely extractable “polars” were multi component and each were < 5 % applied radioactivity (AR)..

b) “Others” consists of multiple components and includes all peaks not quantified in the table above; HPLC and/or TLC chromatograms demonstrate that multi component “others” were < 5 % AR.

ND - Not Detected or < 0.1 %AR; % metabolite water = (% by HPLC in water) × (% of applied radioactivity in the water fraction)

Note: All reported values are rounded. All calculations are based on unrounded values.

  

Table:Distribution of [thiazole-2-14C]-test substance in the Total Water/Sediment Test System of the Taunton River Aerobic Test System (values as percentage of Applied Radioactivity)

[thiazole-2-14C]-test substance

Replicate

Sampling time (DAT)

0

7

14

30

48

70

100

Parent compound

A

93.76

81.88

65.75

23.01

26.50

23.25

13.63

B

95.26

77.69

65.69

33.11

19.04

27.26

13.19

Mean

94.51

79.79

65.72

28.06

22.77

25.26

13.41

Polar(a)

A

ND

1.13

7.72

5.47

1.58

0.42

ND

B

ND

1.27

7.13

1.58

2.51

ND

0.36

Mean

ND

1.20

7.43

3.52

2.00

0.21

0.18

Unspecified metabolite

A

ND

ND

ND

ND

ND

ND

ND

B

ND

ND

ND

ND

ND

ND

ND

Mean

ND

ND

ND

ND

ND

ND

ND

M3

A

ND

3.55

2.99

11.97

10.68

17.25

13.72

B

ND

2.67

2.48

9.95

12.92

16.82

13.86

Mean

ND

3.11

2.74

10.96

11.80

17.04

13.79

M2

A

ND

ND

1.14

2.93

8.00

1.94

1.10

B

ND

1.73

1.05

1.62

5.68

1.94

0.25

Mean

ND

0.87

1.10

2.28

6.84

1.94

0.67

M1

A

ND

0.89

2.79

4.13

6.40

4.87

2.47

B

ND

1.40

1.79

4.47

5.63

4.20

2.08

Mean

ND

1.14

2.29

4.30

6.01

4.54

2.27

Others(b)

A

ND

ND

3.65

2.77

ND

1.41

1.33

B

ND

ND

6.48

ND

ND

0.54

1.01

Mean

ND

ND

5.07

1.37

ND

0.98

1.17

a) HPLC and/or TLC chromatograms demonstrate that freely extractable “polars” were multi component and each were < 5 % applied radioactivity (AR)..

b) “Others” consists of multiple components and includes all peaks not quantified in the table above; HPLC and/or TLC chromatograms demonstrate that multi component “others” were < 5 % AR.

ND - Not Detected or < 0.1 %AR; % metabolite water = (% by HPLC in water) × (% of applied radioactivity in the water fraction)

Note: All reported values are rounded. All calculations are based on unrounded values. 

 

Table: Summary of Extractability and metabolite distribution by soft extraction and harsh extraction (values as percentage of Applied Radioactivity)

Day

Radiolabelled position

PES

Soft extraction

Harsh extraction

Test substance

Polars*

M3

Unknown

100

Oxadiazine

46.48

4.04

10.09

0.48

4.67

8.11

0.86

100

Thiazole

47.88

5.09

8.95

0.32

8.75

3.57

1.41

*“polars” were multi component and each were < 5 % applied radioactivity (AR).

 

Table: Summary of Recovery of Radioactivity by Further Extraction and Humus Fractionation Expressed (values as percentage of Applied Radioactivity)

Day

Radiolabelled position

PES

Soft extraction

Harsh extraction

Fulvic Acid

Humic Acid

Humin

100

Oxadiazine

46.48

4.04

10.1

10.35

3.85

18.15

100

Thiazole

47.88

5.09

8.95

10.19

4.18

19.47

 

Table: Summary of DT50, DT90 and Rate Constants for the Water Phase of the Test Systems during Aerobic Transformation of [14C]-test substance

Radiolabel

SFO

Rate of Dissipation from water phase

Rate of Degradation in whole system

DT50 (days)

DT90 (days)

Χ2

DegT50 (days)

DegT90 (days)

Χ2

[Oxadiazine-4-14C]

19.2

63.8

9.034

29.2

97.9

6.35

[Thiazole-2-14C]

14.8

49.1

11.31

25.2

83.6

12.35

Validity criteria fulfilled:
yes
Conclusions:
The transformation of [oxadiazine-4-14C]-test substance and [thiazole-2-14C]-test substance in aerobic aquatic sediments was assessed in one test system (water pH characteristics < pH 7) that was continuously aerated in the dark at a temperature of 20 ± 2°C for 100 days. The overall mass balance for the [oxadiazine-4-14C]-test substance and [thiazole-2-14C]-test substance in the Taunton River test system was 95.8 % and 97.9 % AR, during the 100-day study, which satisfied the guideline requirement of maintaining material balance between 90 % and 110 % AR. The test substance was shown to partition from the water phase to the sediment with dissipation half-lives ranging from 15 to 19 days. Transformation of the test substance in the water phase and sediment resulted in degradation half-lives ranging from 25 to 29 days and the formation of four products, including CO2. Three components were identified as M3, M2 and M1 by co-chromatography using an HPLC and TLC system. In addition, two metabolites (M3 and M1) were confirmed by LC/MS/MS. The remaining unidentified components comprised seven minor peaks, with no single unknown peak greater than 5 % AR. Further degradation resulted in formation of polar products (in bound residues), mineralization and strong binding to the humin fraction of the sediment organic matter. The bound residues were composed of non-discrete polar residues with no single metabolite > 5 %. Additional levels of the test substance were detected but were present < 0.63 % AR.
Executive summary:

The biodegradation of the test substance in water/sediments system was investigated in a study according to OECD TG 308 in compliance with GLP criteria. In this study, the rate and route of degradation of [14C]-test substance, labelled in the oxadiazine or thiazole ring, was investigated in one aquatic sediment and associated overlaying water collected from Taunton River (Sandy Loam). Water was treated with [oxadizine-4-14C]-test substance or [thiazole-2-14C]-test substance to achieve target concentrations of 0.1 mg/L in the water phase (based on the maximum single label application rate of 300 g a.s./ha). The soils were incubated under aerobic conditions in the laboratory and maintained in dark conditions at 20°C for up to 100 days. Duplicate samples were taken for analysis at seven time intervals. Average mass balance for [oxadiazine-4-14C]-test substance ranged from 92.8 to 105 % of the applied radioactivity (AR). Similarly, average material balance for [thiazole-2-14C]-test substance ranged from 93.1 to 100 % over the course of the 100-day study. Half-lives (DT50) for the dissipation of [oxadiazine-4-14C]-test substance and [thiazole‑2-14C]-test substance in the water phase averaged 19.2 and 14.8 days, respectively. The half-lives of the degradation of [oxadiazine-4-14C]-test substance and [thiazole‑2-14C]‑test substance in the total aquatic sediment system averaged from 29.2 and 25.2 days, respectively. The DegT50 and DegT90 values were calculated using non-linear regression and first-order kinetics (SFO). The 14CO2 evolution due to mineralization of [oxadiazine-4-14C]-test substance ranged from 0.09 to 7.19% AR. A slightly greater degree of mineralization was observed in the test system dosed with [thiazole-2-14C]-test substance as 0.17 and 11.96 % AR were trapped. Negligible quantities of radioactivity, i.e., ≤ 0.6 % AR, were detected in the volatile organic compound (VOC) traps in the systems treated with either [oxadiazine-4-14C]-test substance or [thiazole‑2-14C]‑test substance. Transformation products of[oxadiazine-4-14C]-test substance and [thiazole‑2-14C] test substance were observed in the aquatic aerobic sediment systems as evidenced by three major and several minor peaks in the HPLC profiling with three transformation products being observed consistently. Three products had HPLC retention times of 17, 30, and 33 min (labelled as according to the original HPLC profiling) and were identified as M3, M2 and M1, respectively, by HPLC and TLC co‑chromatography with authentic reference standards. Products M3 and M1 were confirmed by LC/MS/MS. Average non-extractable residues increased to levels of 48.8 % and 51.3 % AR at 100 days after treatment in the test systems treated with [oxadiazine-4-14C]-test substance and [thiazole-2-14C]-test substance, respectively. Extensive efforts were made to further characterize the non-extractable residues (PES). No significant levels of test substance (< 0.63 % AR) were detected in either the “soft” or the “harsh” (reflux) extracts, only polar non-discrete residues at levels < 8.75 % AR and M3 were present at 8.11 % AR. Radioactivity remaining in the PES samples after soft and harsh extractions, was characterized by fractionation into humin, humic acid and fulvic acid fractions in which the majority of the bound radioactivity was associated with the humin fraction (i.e., approximately 18.2 % to 19.5 % AR).

Description of key information

All available data were assessed and the studies representing the worst-case effects were included as key or weight-of-evidence studies. Other studies are included as supporting information. The key studies are considered to be worst-case and were selected for the CSA.

DT50 (freshwater) = 19.2 days; DT50 (system) = 29.2 days, under realistic environmental conditions, OECD 308, Kang 2015.

Key value for chemical safety assessment

Half-life in freshwater:
19.2 d
at the temperature of:
20 °C
Half-life in freshwater sediment:
29.2 d
at the temperature of:
20 °C

Additional information

Table: Overview of available data on biodegradation in water and sediment

Compartment

Radiolabel

Guideline / GLP

Endpoint

Value

Comment

Reference

Natural water: freshwater

[Thiazole-2-14C]-labelled

OECD 308/GLP

DT50

14.8 days

- The 14CO2 evolution due to mineralization of [oxadiazine-4-14C]-test substance ranged from 0.09 to 7.19% AR. A slightly greater degree of mineralization was observed in the test system dosed with [thiazole-2-14C]-test substance as 0.17 and 11.96 % AR were trapped.

- Metabolites M1, M2 and M3 were identified

Kang, 2015

Natural water: freshwater

[Oxadiazine-4-14C]-labelled

DT50

19.2 days

Natural water / sediment: freshwater

[Thiazole-2-14C]-labelled

DT50

25.2 days

Natural water / sediment: freshwater

[Oxadiazine-4-14C]-labelled

DT50

29.2 days