Pirimiphos-methyl

Biodegradation in soil

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in soil: simulation testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Not specified

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

according to guideline

Guideline:

other: according to in-house guidelines (Hill, 1974)

Deviations:

not specified

Principles of method if other than guideline:

The degradation of radiolabelled test substance was studied in four different soils incubated under aerobic conditions. Both non-sterile and sterile soils (via γ irradiation or autoclave) were used. The study was carried out in 19 ± 1˚C darkness for 30 weeks. Volatile and gaseous products from pesticide degradation were 'trapped' and radioactivity in soils was extracted and analysed. The degradation rate and the transformed products of the test substance in soil was studied.

GLP compliance:

no

Remarks:

The study was performed pre-date GLP principles

Test type:

laboratory

Radiolabelling:

yes

Oxygen conditions:

aerobic/anaerobic

Soil classification:

USDA (US Department of Agriculture)

Year:

1976

Soil no.:

#1

Soil type:

sandy loam

% Clay:

19

% Silt:

24

% Sand:

57

% Org. C:

3.65

pH:

7.1

CEC:

19 meq/100 g soil d.w.

Soil no.:

#2

Soil type:

loam

% Clay:

35

% Silt:

17

% Sand:

48

% Org. C:

5.97

pH:

7.5

CEC:

29 meq/100 g soil d.w.

Soil no.:

#3

Soil type:

other: 'Peat'

% Clay:

5

% Silt:

21

% Sand:

74

% Org. C:

36.2

pH:

6

CEC:

92.3 meq/100 g soil d.w.

Soil no.:

#4

Soil type:

sandy loam

% Clay:

7

% Silt:

20

% Sand:

73

% Org. C:

1.04

pH:

6.6

CEC:

5 meq/100 g soil d.w.

Details on soil characteristics:

SOIL COLLECTION AND STORAGE
- Geographic location: See Table 1 in 'Any other information on materials and methods incl. tables'.
- Collection procedures: Using a clean spade or trowel several randomly distributed sub samples were taken.
- Sampling depth (cm): No greater than 10 cm
- Transportation: The collected soil was transported to the laboratory in thin polythene bags to help preserve moist, aerobic conditions (polyethylene allows the passage of air but not water vapour) and reduce any tendency for fluctuations in the microbial populations. All soils were returned to the laboratory within 24 hours of sampling (soils held overnight being left in a cool place with the top of the bag loosely folded over).
- Soil preparation: Soils were prepared for incubation with the minimum of delay (experiments were started within 3-4 days of receipt of the soil in the laboratory). Each soil was thoroughly mixed and passed through a 3 mm sieve (to remove root material, stones, etc) and thoroughly mixed by gentle rotation in a closed container. Any soil too wet to sieve was air dried, at laboratory temperature, but only to point whereby sieving was facilitated.
Prior to treatment with pesticide, and commencement of incubation, water was added to the soil (by mixing deionised water with bulk sieved soil) to a level such that the final treatment with pesticide results in a soil with moisture level of 40% of MHC (30% for peat soils, as at 40% of MHC such soils are too wet to handle satisfactorily). Soils were sterilised by either 5 megarads gamma radiation or autoclaving.

PROPERTIES OF THE SOILS: See Table 2 in 'Any other information on materials and methods incl. tables'.

Duration:

30 wk

Soil No.:

#1

Initial conc.:

1 kg/ha d.w.

Based on:

act. ingr.

Remarks:

Measured application rate: 0.7 kg/ha d.w.

Soil No.:

#1

Initial conc.:

2 kg/ha d.w.

Based on:

act. ingr.

Remarks:

Measured application rate: 2.0 kg/ha d.w.

Soil No.:

#1

Initial conc.:

10 kg/ha d.w.

Based on:

act. ingr.

Remarks:

Measured application rate: 8.7 kg/ha d.w.

Soil No.:

#2

Initial conc.:

1 kg/ha d.w.

Based on:

act. ingr.

Remarks:

Measured application rate: 0.7 kg/ha d.w.

Soil No.:

#2

Initial conc.:

2 kg/ha d.w.

Based on:

act. ingr.

Remarks:

Measured application rate: 2.0 kg/ha d.w.

Soil No.:

#2

Initial conc.:

10 kg/ha d.w.

Based on:

act. ingr.

Remarks:

Measured application rate: 8.7 kg/ha d.w.

Soil No.:

#3

Initial conc.:

1 kg/ha d.w.

Based on:

act. ingr.

Remarks:

Measured application rate: 0.7 kg/ha d.w.

Soil No.:

#4

Initial conc.:

1 kg/ha d.w.

Based on:

act. ingr.

Remarks:

Measured application rate: 0.7 kg/ha d.w.

Parameter followed for biodegradation estimation:

CO2 evolution

radiochem. meas.

Soil No.:

#1

Temp.:

19 ± 1 ˚C

Humidity:

40% MHC

Soil No.:

#2

Temp.:

19 ± 1 ˚C

Humidity:

40% MHC

Soil No.:

#3

Temp.:

19 ± 1 ˚C

Humidity:

30% MHC

Soil No.:

#4

Temp.:

19 ± 1 ˚C

Humidity:

40% MHC

Details on experimental conditions:

EXPERIMENTAL DESIGN (An overview of the experimental design is provided in Table 3 in 'Any other information on materials and methods incl. tables' )
- Soil: 30 g (except 20 g ‘Blackborough', a peat )
- Soil incubation conditions: The soils were incubated in glass crystallising dishes (4cm diam, 3 cm high), supported on siliconised wire racks inside glass 'cylinders', with a stream of either CO2-free air or oxygen-free nitrogen gas. In addition to incubation in atmospheres of air and nitrogen, pesticide degradation studies were carried out under waterlogged conditions (flooding the soil 6 hrs after pesticide application). The effect of waterlogging and non-waterlogged anaerobiosis on the degradation of the pesticide and its breakdown products, after a period or aerobic soil incubation, was also studied. Pots of soil were removed from 'Gore' and 'Peartree 7' aerobic incubation units 5 weeks after pesticide application, and subsequently either incubated in an atmosphere of nitrogen, or waterlogged. For waterlogged soils, 6cm high pots are used. Addition of sterile deionised water (2.5 cm depth) was added to above the soil surface.
- No. of replication treatments: 1
- Efluent air traps: Effluent air or nitrogen gas from the soil incubation units was passed through a series of traps containing, in sequence, 0.05M H2SO4, 2-methoxyethanol, ethanolarnine (two tubes), to 'trap' volatile and gaseous products of pesticide degradation.
- Identity and concentration of co-solvent:

TEST MATERIAL APPLICATION
Both biologically active and sterilised soils were treated with radiolabelled test substance. Immediately prior to treatment of biologically active (non-sterile) soils with the pesticide, pre-determined amounts of unlabelled test substance were added to separate preparations of the labelled pesticide in ethyl acetate. Aqueous preparations of the pesticide, for treatment of soils, were made by the addition of water and subsequent removal of the solvent by rotary evaporation, under vacuum at 30˚C. 0.5 mL aliquots of the aqueous 14C preparations were added to 4 cm diameter pots containing 30 g soil (20 g ‘Blackborough', a peat). The concentrations of the preparations were such that each pot should have received a surface application of either 0.126 mg or 1.26 mg pesticide (equivalent to 1 kg/ha or 10 kg/ha active ingredient respectively), with the level of applied radioactivity at 1-3 μCi per pot of soil. Sterile soils were treated at the rate of 2 kg/ha (0.252mg pesticide). 'Gore' and 'Peartree 7', sterilised by Y-radiation and by autoclaving, were treated with sterile 14C labelled test substance and sterility maintained throughout the incubation period (confirmed by incubation of samples of soil in nutrient broths and on agar.
The amount of radioactivity per unit volume of the aqueous preparations was found to vary considerably during the period required for application of the pesticide to the surface of the soil in pots. This was due to the low water solubility of the pesticide resulting in 'loss' of the chemical onto the glass surfaces in which the preparations were held. The aqueous pesticide preparations were cloudy and pesticide 'lost' from solution during application could later be recovered from the glass vessel walls with acetone. It was subsequently found that these losses could be avoided by addition of methanol to the aqueous preparations, at a final concentration of 10%.
Treatment of sterile soils was carried out at a later date than the non-sterile soils, therefore aqueous pesticide preparations, containing 10% methanol, were added to sterile soils. The soils used for sterilizing were the same as used for non-sterile incubation and were sterilized at the start of the non-sterile studies.
Radioactive test substance was sterilized by membrane filtration of the methanolic solutions, immediately prior to addition of sterile distilled
water to form the aqueous pesticide preparation. All preparatory work and soil treatments were carried out in a laminar flow sterile cabinet, using sterile apparatus and aseptic techniques.

EXPERIMENTAL CONDITIONS
- Moisture maintenance method: Moisture levels in the soils are maintained by the addition of sterile deionised water, approximately every 10 weeks, to restore the pot of soil to its original weight.
- Continuous darkness: Yes

SAMPLING DETAILS
- Sampling intervals: The amount of radioactivity added to each soil pot was determined by scintillation counting samples of the aqueous pesticide preparations. Samples were taken both before and after addition of the pesticide to soil to check for losses of radioactivity in solution, due to volatilization, adsorption on to glass surfaces, etc..
Complete pots of soil were removed from the incubation units for analysis 2 hours after application of the pesticide to the soil surfaces, after that samples were taken in week 2, 5, 10 and 30 after pesticide application. Where soils were waterlogged, the samples were taken 6 hrs
after pesticide application, and then in 12 hrs, 2, 5, 10 and 30 weeks later.
- Sampling method for soil samples: Sterile soil was sampled in laminar flow cabinets using aseptic techniques and sterile instruments. To maintain an anaerobic environment in the soils incubated in an atmosphere of nitrogen, sampling, watering etc was carried out in an inflatable hood flushed several times with nitrogen (after addition of apparatus, instruments and sealed ‘soil cylinder'. The sterility of the sampled sterile soils was checked by addition of a soil suspension to both broth and agar media.

Soil No.:

#1

% Recovery:

79

Remarks on result:

other: Aerobic; 1 kg/ha exposure rate

Soil No.:

#1

% Recovery:

75

Remarks on result:

other: Aerobic; 10 kg/ha exposure rate

Soil No.:

#2

% Recovery:

88

Remarks on result:

other: Aerobic; 1 kg/ha exposure rate

Soil No.:

#2

% Recovery:

65

Remarks on result:

other: Aerobic; 10 kg/ha exposure rate

Soil No.:

#3

% Recovery:

77

Remarks on result:

other: Aerobic; 1 kg/ha exposure rate

Soil No.:

#4

% Recovery:

100

Remarks on result:

other: Aerobic; 1 kg/ha exposure rate

Parent/product:

parent

Soil No.:

#1

% Degr.:

99.47

Parameter:

radiochem. meas.

Sampling time:

30 wk

Remarks on result:

other: Aerobic; 1 kg/ha exposure rate

Parent/product:

parent

Soil No.:

#1

% Degr.:

99.7

Parameter:

radiochem. meas.

Sampling time:

30 wk

Remarks on result:

other: Aerobic; 10 kg/ha exposure rate

Parent/product:

parent

Soil No.:

#2

% Degr.:

99.38

Parameter:

radiochem. meas.

Sampling time:

30 wk

Remarks on result:

other: Aerobic; 1 kg/ha exposure rate

Parent/product:

parent

Soil No.:

#2

% Degr.:

99.96

Parameter:

radiochem. meas.

Sampling time:

30 wk

Remarks on result:

other: Aerobic; 10 kg/ha exposure rate

Parent/product:

parent

Soil No.:

#3

% Degr.:

99.61

Parameter:

radiochem. meas.

Sampling time:

30 wk

Remarks on result:

other: Aerobic; 1 kg/ha exposure rate

Parent/product:

parent

Soil No.:

#4

% Degr.:

99.98

Parameter:

radiochem. meas.

Sampling time:

30 wk

Remarks on result:

other: Aerobic; 1 kg/ha exposure rate

Key result

Soil No.:

#1

DT50:

11.1 d

Type:

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

Temp.:

19 °C

Remarks on result:

other: Recalculated based on extracted test substance; aerobic; 1 kg/ha

Key result

Soil No.:

#1

DT50:

14.6 d

Type:

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

Temp.:

19 °C

Remarks on result:

other: Recalculated based on extracted test substance; aerobic; 10 kg/ha

Key result

Soil No.:

#2

DT50:

5 d

Type:

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

Temp.:

19 °C

Remarks on result:

other: Recalculated based on extracted test substance; aerobic; 1 kg/ha

Key result

Soil No.:

#2

DT50:

10.4 d

Type:

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

Temp.:

19 °C

Remarks on result:

other: Recalculated based on extracted test substance; aerobic; 10 kg/ha

Key result

Soil No.:

#4

DT50:

4.7 d

Type:

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

Temp.:

19 °C

Remarks on result:

other: Recalculated based on extracted test substance; aerobic; 1 kg/ha

Soil No.:

#1

DT50:

0.75 wk

Type:

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

Temp.:

19 °C

Remarks on result:

other: Aerobic; original report; 10 kg/ha

Soil No.:

#1

DT50:

1.5 wk

Type:

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

Temp.:

19 °C

Remarks on result:

other: Aerobic; original report; 1 kg/ha

Soil No.:

#2

DT50:

0.5 wk

Type:

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

Temp.:

19 °C

Remarks on result:

other: Aerobic; original report; 1 kg/ha

Soil No.:

#2

DT50:

0.5 wk

Type:

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

Temp.:

19 °C

Remarks on result:

other: Aerobic; original report; 10 kg/ha

Soil No.:

#3

DT50:

0.5 wk

Type:

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

Temp.:

19 °C

Remarks on result:

other: Aerobic; original report; 1kg/ha

Soil No.:

#4

DT50:

0.5 wk

Type:

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

Temp.:

19 °C

Remarks on result:

other: Aerobic; original report; 1 kg/ha

Transformation products:

not specified

Remarks:

See additional information provided in overall endpoint summary.

Details on transformation products:

An overview of the results in provided in Table 6 - Table 10 in 'Any other information on results incl. tables'.

EXTRACTABLE DEGRADATION PRODUCTS
- Non-sterile aerobic soils: In all four soils the major product identified was M1 derived from the hydrolysis of the phosphorothioate moiety from the pyrimidine ring. The amounts of M1 increased to between 36% and 56% of the total recovered radioactivity from 'Peartree 7' and 'Frensham' respectively after 2 weeks incubation of the test substance in soil. The amounts of M1 decreased in all soils during the remainder of the incubation period. In 'Peartree 7' and 'Gore' incubated with 10 kg/ha test substance the proportions of M1 formed were similar to those in soils treated at the 1 kg/ha rate. Small amounts of M2, and M3 were also extracted, each amounting to 4% or less of the total recovered radioactivity. M5 was observed on autoradiographs but was not separately quantified. Product 'z' which chromatographed with the test substance on TLC's developed in EHM was separated from the parent on TLC's developed in HA and was separately quantified. This product did not cochromatograph with any available marker. Product 'y' appeared as a minor radioactive product on TLC's developed in EHM. This was shown to co-chromatograph with the reference compound M4.

- Non-sterile anaerobic soils ('Peartree 7' and 'Gore'): In both soils incubated under nitrogen, the proportions of radioactive degradation products differed slightly from those in the corresponding aerobic soils. Larger amounts of M1 were present in these soil extracts. Where soils were transferred to anaerobic conditions after 5 weeks aerobic incubation, the rate of breakdown of the test substance was slightly reduced compared with that in the corresponding aerobic soils but less than in those soils incubated anaerobically from the time of application of the test substance.
The rate of degradation and distribution of the test substance degradation products in waterlogged soils was similar to that in the anaerobic soils incubated under nitrogen.

- Sterile soils: Degradation in both autoclaved and γ irradiated soils incubated aerobically was considerably slower than in the corresponding aerobic soils. M1 was again the major degradation product and amounted to approximately 28% of recovered radioactivity in 'Peartree 7' autoclaved and 37% in 'Gore' γ irradiated. M2, M3 and an unknown product were also identified but none amounted to more than approximately 7% of recovered radioactivity.

POLAR DEGRADATION PRODUCTS
Throughout the experimental incubation period, increasing amounts of polar radioactive products were extracted from soils. By week 30 these accounted for between 22% and 51% of recovered radioactivity. Most of these products remained on or near the baseline of chromatograms developed in EHM. In acetone extracts of soils treated with the test substance, the majority of the polar radioactivity was shown to be N,N,diethylguanidine.
- Chromatographic identification of polar products: All the polar radioactivity, in the acetone extracts, which remained on the baseline of TLC plates developed in EHM was satisfactorily resolved in BEW. Despite the diffuse nature of the polar radioactivity in this solvent the only marker to co-chromatograph with the polar radioactivity was N,N, diethylguanidine (DEG).

- Reverse isotope dilution: Results from reverse isotope dilution studies showed that the proportions of radioactivity present as DEG were very similar to those obtained from chromatography. The specific activities of the re-crystallised DEG samples remained approximately the same after 5 - 7 re-crystallisations. With samples of re-crystallised EG, and G the specific activities dropped to zero after one and four re-crystallisations respectively. These results indicate that there was no EG or G present in the soil extracts.

Evaporation of parent compound:

not specified

Volatile metabolites:

yes

Residues:

yes

Details on results:

An overview of the results in provided in Table 4 - Table 11 in 'Any other information on results incl. tables'.

- Recovery of Radioactivity from Soils: The amounts recovered ranged from 61% to 176% of that applied, but the majority ranged from 84% to 113% of applied radioactivity. There was no consistent decrease or increase in the amounts of radioactivity recovered with increasing time of soil incubation. The variable recoveries were probably due to the effects of loss of the pesticide on to glass surfaces during its application and to its low solubility in water.

- Gaseous and volatile products from soil: The only 'trapping' solution to contain radioactivity during the incubation period was the first ethanolamine ‘trap’. This radioactivity has been identified as 14CO2.
- 14CO2 from non-sterile aerobically incubated soils: 14CO2 was evolved from all non-sterile aerobic soils. 14CO2 evolution occurred most rapidly from 'Gore' at approximately twice the rate observed in ‘Peartree 7', 4 times that in 'Frensham' and 9 times that in 'Blackborough'. After 30 weeks incubation of 'Gore'. with 1 kg/ha test substance, 17% of the applied pesticide had been evolved as 14CO2; 7% of pesticide applied at 10 kg/ha was evolved from 'Gore' in the same period. A similar reduction was observed in the 'Peartree 7' 10 kg/ha treatment. Cleavage of the pesticide pyrimidine ring thus occurred in all non-sterile soils incubated aerobically.

- 14CO2 from non-sterile anaerobic soils (‘Gore’ and ‘Peartree 7’ only)
1) Using Nitrogen gas: The rate of 14CO2 evolution was substantially reduced in soils incubated anaerobically in a stream of nitrogen gas, compared with that from soils incubated aerobically. Less than 2% was evolved from 'Gore’ and less than 1% from 'Peartree 7' after 30 weeks incubation. Where 'Gore’ and 'Peartree 7’ soils had been transferred to an atmosphere of nitrogen after 5 weeks aerobic incubation, the rates of 14CO2 evolution were reduced. They were similar to those exhibited by the permanently anaerobic soils.
2) Water logging: Waterlogging ‘Gore’ and ‘Peartree 7’ soils from week 0 markedly reduced the rate of 14CO2 evolution. In ‘Gore’ there was a 3-fold reduction in the rate of 14CO2 evolution compared with that from the corresponding aerobically incubated soils; and in 'Peartree 7’ the rate was 6-fold lower. The effect of waterlogging 'Gore' and ‘Peartree 7' after 5 weeks aerobic incubation was to reduce the rate of 14CO2 evolution to approximately that shown by the permanently waterlogged soils.

- 14CO2 from aerobically incubated sterile soils: Less than 0.3% 14CO2 was evolved from γ-irradiated 'Gore' soil and less than 0.1% from ‘Peartree 7’ after 20 weeks incubation. No measurable 14CO2 was evolved from either of the autoclaved soils during the incubation period of 10 weeks. The small amount of 14CO2 from γ-irradiated ‘Gore' and ‘Peartree 7' could indicate the presence of biological activity in these soils, such as functional enzymes which are destroyed by autoclaving.

- Extraction of radioactivity from soils: In the majority of non-sterile soils, > 99% of radioactivity present (as a percentage of that recovered by extraction, and combustion of extracted soil) was recovered at zero time by soxhlet extraction with acetone. After 30 weeks soil incubation, soxhlet extraction in acetone recovered between 20% and 51% of the radioactivity remaining in the soil.
Refluxing the acetone extracted soils with methanol : water (1:1) recovered between 10% and 36% more radioactivity. The amounts of non-extractable, 'bound', radioactivity from combustion of solvent extracted soils ranged from 28% to 66% by 30 weeks incubation. The highest amount of bound radioactivity was in 'Blackborough', (incubated aerobically), a fen peat soil with a high organic matter content. In general, the proportions of 'bound' radioactivity increased with increasing time of incubation. In sterile soils (γ-irradiated) similar increases in binding occurred out to a maximum of 32% by week 30 in 'Peartree 7'. In 'Gore' and 'Peartree 7' autoclaved soils, after '10 weeks incubation, less than 5% of recovered radioactivity was 'bound'.

- Residual extractable test substance in soils: The test substance was rapidly degraded in all non-sterile soils. After 10 weeks incubation, less than 4% of the test substance could be recovered in soil extracts. incubation of the test substance in 'Gore' and 'Peartree 7' soils under waterlogged or anaerobic (N2) conditions using nitrogen, did not noticeably affect the rate of degradation of the parent pesticide.
In all sterile soils the rate of degradation of the pesticide was much reduced. After 10 weeks incubation approximately 32% - 60% of the parent pesticide was present in both γ irradiated and Autoclaved soils. In all non-sterile soils less than 50% of the parent pesticide remained in soil after 1.5 weeks incubation and less than 10% after 7.5 weeks. In the sterile soils the times taken for degradation of 50% of the parent pesticide range from approximately 5 - 16 weeks. Degradation of 90% of the pesticide did not occur in sterile soils during the incubation period.

Table 4. Recovery of radioactivity from soils treated with 14C labelled test substance

Soil treatment code Recovery of radioactivity (total 14C from extraction of soil, combustion of extracted soil and from 'trapping' solutions) as a percentage of radioactivity added to soil ‘Peartree 7' ‘Gore' ‘Blackborough' ‘Frensham' Soil analysis times (weeks after pesticide application) 0 2 5 10 30 0 2 5 10 30 0 2 5 10 30 0 2 5 10 30 A/1 103 112 120 118 79 103 117 139 83 88 107 95 89 100 77 86 109 79 111 105 A/10 100 84 69 75 100 87 - 77 65 N/1 176 150 - 134 93 90 84 - 80 74 W/1 61 95 - 70 87 104 124 - 121 129 AN/1 - - - 99 101 - - - 98 101 AW/1 - - - 96 106 - - - 99 104 A/(Sγ)2 118 - 113 109 106 112 - 89 113 - A/(SA)2 99 - 88 90 - 99 - 69 91 -

  Table 5. Radioactivity in extracts of soil treated with the test substance

Soil treatment code	Radioactivity in extracts of soil treated with 14C labelled test substance, as a % of recovered radioactivity remaining in soil
	‘Peartree 7'					‘Gore'					‘Blackborough'					‘Frensham'
	Soil analysis times (weeks after pesticide application)
	0	2	5	10	30	0	2	5	10	30	0	2	5	10	30	0	2	5	10	30
A/1	99	82	71	59 (16)	28 (27)	100	78	67	53 (16)	41 (16)	100	89	68	45 (10)	23 (10)	99	83	65	54 (25)	46 (20)
A/10	99	90	-	62 (16)	39 (23)	99	93	-	60 (15)	48 (21)
N/1	98	91	-	73 (11)	26 (23)	98	91	-	66 (6)	34 (14)
W/1	79	87	-	63 (17)	51 (20)	98	85	-	78 (7)	20 (19)
AN/1	-	-	-	59 (15)	44 (21)	-	-	-	46 (18)	49 (10)
AW/1	-	-	-	59 (15)	22 (36)	-	-	-	52 (20)	35 (20)
A/(Sγ)2	99	-	90 (2)	84 (5)	67 (11)	99	-	86 (4)	77 (5)
A/(SA)2	99 (0.5)	-	88 (5)	80 (8)	99 (0.7)	-	92 (2)	85 (4)

Table 6. Distribution of radioactivity from aerobic incubation of 14C labelled test substance in ‘Gore’ and 'Peartree 7' soils.

Soil		Distribution of radioactivity (as a percentage of total recovered 14C from 'trapping' solutions, extraction of a single pot of soil and combustion of extracted soil)
		'Gore' (1 kg/ha)					'Gore' (10 kg/ha)					'Peartree 7' (1 kg/ha)^b						'Peratree 7' (10 kg/ha)
		Weeks incubation
		0	2	5	10	30	0	2	10	30	0		2	5	10	30	0		2	10	30
The test substance and degradation products in extracts of soil^a	Parent substance	71.28	10.07	0.04	0.21	0.62	78.07	30.44	2.04	0.04	78.5		37.7	2.4	0.65	(0.53)	86.6		44.0	4.2	0.3
	M2	3.11	1.37	3.2	0.93	<0.01	2.43	0.93	1.01	0.26	2.08		0.76	4.02	2.65	(0.69)	0.81		1.1	2.55	3.07
	M1	12.10	37.22	15.27	7.82	1.75	6.68	46.70	22.85	1.45	11·40		36.33	40.48	42.08	(6.14)	6.82		36.48	38.02	20.19
	M3	0.33	2.44	1.36	1.73	0.3	0.48	2.04	1.25	0.60	0.09		1.24	1.60	1.10	(0.74)	0.16		0.99	3.16	1.84
	M4	4.16	0.42	1.79	0.78	0.11	0.29	0.36	0.57	0.49	1.00		0.25	1.27	0.62	(1.22)	0.94		0.07	1.12	0.93
	Unknown	3.58	0.54	1.38	0.35	0.11	1.95	0.62	0.43	0.32	0.79		0.24	0.64	0.33	(0.07)	0.50		1.10	1.43	0.57
	remainder	4.38	24.92	40.72	49.75	43.55	9.47	11.37	43.65	51.24	4.87		5.82	19.7	25.4	(17.57)	3.52		6.13	27.27	31.46
14CO2		-	1.70	3.79	10.87	18.34	-	0.58	3.76	10.11	-		0.52	1.01	1.77	4.80	-		0.34	1.33	3.12
'Bound' (non-extractable radioactivity)		1.05	21.4	31.67	27.5	35.30	0.65	6.9	24.47	35.50	1.2		17.16	29	25.38	41.80	0.63		9.86	20.91	37.93

a. M5 was not separately quantified and is included in the 'Remainder' of radioactivity.

b. Distribution of radioactivity in 30 week acetone extract of soil only. Methanol extract 'lost’ during experimental handling.

Table 7.Distribution of radioactivity from aerobic incubation of 14C labelled test substance in ‘Frensham’ and 'Blackborough' soils.

Soil		Distribution of radioactivity (as a percentage of total recovered 14C from 'trapping' solutions, extraction of a single pot of soil and combustion of extracted soil)
		'Frensham' (1 kg/ha)					'Blackborough' (1 kg/ha)
		Weeks incubation
		0	2	5	10	30	0	2	5	10	30
The test substance and degradation products in extracts of soil	Parent substance	85.21	10.87	0.41	1.02	0.12	75.19	18.5	10.05	1.91	0.39
	M2	1.18	0.77	2.60	3.64	2.49	2.06	1.17	0.44	1.02	1.01
	M1	6.40	55.58	36.79	30.86	12.21	8.01	50.75	45.00	33.57	6.49
	M3	0.21	3.02	1.60	1.66	2.02	0.54	1.64	1.69	1.10	0.61
	M4	0.20	0.20	0.43	0.89	0.82	1.06	1.06	< 0.10	0.27	0.78
	Unknown	2.11	0.65	0.34	0.31	0.12	3.2	0.80	1.50	0.54	0.02
	remainder	3.71	11.57	22.50	39.88	45.71	9.47	14.98	9.55	16.09	22.35
14CO2		-	0.57	1.57	1.91	3.67	-	0.30	0.53	0.77	2.09
'Bound' (non-extractable radioactivity)		0.98	16.77	33.74	19.87	32.78	0.48	10.78	31.23	44.71	66.28

Table 8.Distribution of radioactivity from anaerobic incubation of 14C labelled test substance in N2 or waterlogged anaerobic ‘Gore’ soil.

Soil		Distribution of radioactivity (as a percentage of total recovered 14C from 'trapping' solutions, extraction of a single pot of soil and combustion of extracted soil)
		Anaerobic (N2) from zero time				Anaerobic (N2) from week 5		Anaerobic (waterlogged) from zero time				Anaerobic (waterlogged) from week 5
		'Gore' soil; Weeks incubation
		0	2	10	30	10	30	0	2	10	30	10	30
In extracts of soil	Parent substance	57.84	20.8	0.69	0.31	0.03	0.15	74.93	6.42	0.40	0.26	0.60	0.31
	M2	3.35	0.85	1.08	0.63	0.80	0.41	1.05	1.46	2.51	0.90	1.08	<0.01
	M1	26.87	62.55	53.10	13.79	36.04	4.70	15.66	63.67	60.09	3.31	14.30	4.41
	M3	2.24	1.42	1.15	0.28	1.30	0.83	0.70	2.59	2.27	2.25	1.08	1.40
	M4	1.03	0.6	2.14	2.45	0.67	0.96	0.08	0.7	0.48	0.61	1.67	0.68
	Unknown	0.29	0.38	0.81	0.32	0.08	0.15	0.37	0.35	0.31	<0.01	0.25	0.45
	remainder	5.90	4.04	12.21	29.02	20.70	47.08	5.61	9.57	16.69	30.09	43.82	42.46
14CO2		-	0.22	0.44	2.71	6.34	8.36	-	0.46	0.99	4.04	8.05	9.75
'Bound' (non-extractable radioactivity)		2.47	9.11	28.43	50.48	34.07	37.46	1.60	14.77	16.53	58.37	29.04	40.52

 

  Table 9. Distribution of radioactivity from anaerobic incubation of 14C labelled test substance in N2 or waterlogged anaerobic ‘Peartree 7’ soil. 

Soil		Distribution of radioactivity (as a percentage of total recovered 14C from 'trapping' solutions, extraction of a single pot of soil and combustion of extracted soil)
		Anaerobic (N2) from zero time				Anaerobic (N2) from week 5		Anaerobic (waterlogged) from zero time				Anaerobic (waterlogged) from week 5
		' Peartree 7' soil Weeks incubation
		0	2	10	30	10	30	0	2	10	30	10	30
In extracts of soil	Parent substance	82.00	17.14	1.63	0.49	1.86	0.88	51.10	10.41	1.21	0.48	1.25	0.07
	M2	1.15	0.10	0.51	0.14	2.18	0.76	0.93	0.53	2.74	0.48	1.91	1.73
	M1	8.37	63.49	69.57	35.43	40.88	38.00	16.00	61.4	50.00	42.52	45.10	29.76
	M3	0.33	1.01	2.66	1.38	1.09	2.12	0.40	1.23	1.85	0.71	1.05	1.95
	M4	0.29	0.34	0.66	0.36	2.02	0.73	1.34	0.4	2.23	0.26	0.6	0.08
	Unknown	0.76	0.58	1.63	0.13	0.39	0.26	2.17	0.26	0.8	0.49	0.57	0.06
	remainder	4.98	7.03	7.53	9.53	25.31	20.60	7.35	12.54	20.82	25.82	22.50	22.49
14CO2		-	0.11	0.19	0.68	1.46	1.83	-	0.21	0.74	1.06	1.11	2.41
'Bound' (non-extractable radioactivity)		2.11	9.29	15.59	50.74	24.76	34.74	20.7	13.01	19.53	28.2	25.31	41.56

 

 Table 10.Distribution of radioactivity from sterile incubation of 14C labelled test substance in ‘Gore’ and ‘Peartree 7’ soil.

Soil		Distribution of radioactivity (as a percentage of total recovered 14C from 'trapping' solutions, extraction of a single pot of soil and combustion of extracted soil)
		γ irradiated				Autoclaved			γ irradiated				Autoclaved
		'Gore' soil Weeks incubation							'Peartree' soil Weeks incubation
		0	5	10	30	0	5	10	0	5	10	30	0	5	10
In extracts of soil	Parent substance	83.77	21.86	31.49	33.4	86.18	69.27	63.52	71.73	Data not available	57.91	30.75	88.57	54.02	43.13
	M2	2.92	2.13	2.92	1.04	5.85	0.21	1.17	5.64		1.21	3.52	3.19	1.28	1.41
	M1	6.36	37.14	16.44	6.68	2.32	17.31	16.11	11.72		16.34	21.20	4.42	22.01	28.23
	M3	0.75	7.25	7.65	1.36	0.18	0.11	0.90	0.71		1.53	2.49	0.22	1.56	2.10
	M4	1.65	0.18	0.84	0.36	0.27	0.67	0.27	0.37		0.31	0.42	0.63	1.20	1.06
	Unknown	<0.01	0.21	0.64	0.09	0.72	0.64	0.63	1.37		2.6	2.25	0.54	0.26	0.89
	remainder	3.92	21.02	22.29	16.68	4.32	5.59	6.75	7.73		8.82	7.17	2.09	12.70	10.92
14CO2		-	0.05	0.08	0.31	-	<0.01	< 0.01	-	0.01	0.02	0.06	-	<0.01	<0.01
'Bound' (non-extractable radioactivity)		0.64	10.2	17.73	40.02	0.19	6.12	10.62	0.81	7.86	11.36	32.26	0.22	7.0	12.28

 

Table 11. Degradation of 50% and 90% of the test substance in soil

Soil	Soil treatment code	Time taken, in weeks for the degradation of the test substance
		50%	90%
‘Gore'	A/1	0.5	1.5
‘Peartree 7'		0.5	3.5
‘Blackborough'		0.75	5
‘Frensham'		0.5	2
‘Gore'	A/10	0.5	6
‘Peartree 7'		1.5	7.5
‘Gore'	N/1	0.5	5
‘Peartree 7'		0.5	3
‘Gore'	W/1	0.5	2
‘Peartree 7'		0.5	2.5
‘Gore'	AN/1	a	a
‘Peartree 7'		a	a
‘Gore'	AW/1	a	a
‘Peartree 7'		a	a
‘Gore'	A/(Sγ)2	5	NE
‘Peartree 7'		16	NE
‘Gore'	A/(SA)2	15^b	NE
‘Peartree 7'		9	NE

a At the time of waterlogging or transfer of soil to an atmosphere of nitrogen, 50% and 90% of the pesticide had already been degraded.

NE - Not estimated: the incubation period was not long enough for the 90% degradation level to be reached, nor was data sufficient to estimate a value.

b Estimated value.

Conclusions:

Rapid degradation of the test substance in non-sterile soils was observed under aerobic conditions. The DT50 values ranged between 4 and 10 days. Recalculated (normalised) DT50 values ranged between 4.7 to 14.6 days. Small amounts of 14CO2 evolved in γ- irradiated sterile soils, indicating that degradation of the test substance occurred by a biological mechanism. However, the presence of M1 in both autoclaved and γ irradiated soils showed that the test substance was also degraded by chemical hydrolysis.

Executive summary:

The degradation of 14C radiolabelled test substance has been studied in four soils of differing characteristics incubated under aerobic and anaerobic conditions. The study was conducted according to in-house guidelines (Hill, 1974). These methods pre-date GLP principles and official guidelines but meet most of the current guideline’s points and are deemed acceptable for assessment. Both non-sterile and sterile soils (via γ irradiation or autoclave) were used. Anaerobic soils were incubated under nitrogen gas or in waterlogged. The study was carried out in 19 ± 1˚C darkness for 30 weeks. The application rate was 1, 2 and 10 kg/ha. Volatile and gaseous products from degradation were 'trapped' and radioactivity in soils was extracted and analysed. The degradation rate and the transformed products of the test substance in soil was studied.

 

After 30 weeks incubation, the recovered range of the radioactivity from soils were from 61% to 176% of that applied, but the majority ranged from 84% to 113%. There was no consistent decrease or increase in the amounts of radioactivity recovered with increasing time of soil incubation. The only 'trapping' solution to contain radioactivity during the incubation period was the first ethanolamine ‘trap’. This radioactivity has been identified as 14CO2.

The test substance was rapidly degraded in all non-sterile soils. After 10 weeks incubation, less than 4% of the test substance could be recovered in soil extracts. incubation of the test substance in 'Gore' and 'Peartree 7' soils under waterlogged or anaerobic (N2) conditions using nitrogen, did not noticeably affect the rate of degradation of the test substance. In all sterile soils the rate of degradation of the test substance was much reduced. After 10 weeks incubation approximately 32% - 60% of the test substance was present in both γ irradiated and Autoclaved soils. In all non-sterile soils less than 50% of the test substance remained in soil after 1.5 weeks incubation and less than 10% after 7.5 weeks. In the sterile soils the times taken for degradation of 50% of the test substance range from approximately 5 - 16 weeks. Degradation of 90% of the pesticide did not occur in sterile soils during the incubation period.

Degradation of the test substance mainly occurred by loss of the phosphorothioate side chain from the molecule and by opening of the pyrimidine ring as shown by the release of 14CO2, thus the major product from degradation of the test substance in all soils was M1. Minor products M2, M3, M4 and M5 were also formed. Subsequently cleavage of the pyrimidine ring took place with the formation of polar products, the major component of which was N,N,-diethylguanidine. The proportion of 'bound' (non-extractable) radioactivity in soils increased with increasing time of incubation. From 16% to 45% of recovered radioactivity was 'bound' after 10 weeks incubation of the test substance in non-sterile soils. In sterile soils up to 17% and 11% of recovered radioactivity was 'bound' in γ irradiated and autoclaved soils respectively in the same time period.

Based on the findings, the DT50 values ranged between 4 and 10 days. Recalculated (normalised) DT50 values ranged between 4.7 to 14.6 days.

Description of key information

Geometric mean DT50 in soil = 8.3 d, 19 °C, no guideline followed, Arnold 1976

Key value for chemical safety assessment

Half-life in soil:

8.3 d

at the temperature of:

19 °C

Additional information

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

USDA / Name / Origin	OC [%] / pH (water)	T [˚C] / Moisture [%] / Application rate	DT50 [wk] (Kinetic model in original report)	DT50 [d] (Kinetic model recalculated)	Author / year
Sandy loam / Peartree 7 / Berkshire, UK	3.65 / 7.1	19 / 40% MHC / 1 kg/ha	0.75 week (Visual)	11.1 (SFO)	Arnold et al. / 1976
Sandy loam / Peartree 7 / Berkshire, UK	3.65 / 7.1	19 / 40% MHC / 10 kg/ha	1.5 week (Visual)	14.6 (SFO)	Arnold et al. / 1976
Loam / Gore / Bershire, UK	5.97 / 7.5	19/ 40% MHC / 1 kg/ha	0.5 week (Visual)	5.0 (SFO)	Arnold et al. / 1976
Loam / Gore / Bershire, UK	5.97 / 7.5	19/ 40% MHC / 10 kg/ha	0.5 week (Visual)	10.4 (SFO)	Arnold et al. / 1976
Sandy loam / Frensham / Hampshire UK	1.04 / 6.6	19/ 40% MHC / 1 kg/ha	0.5 week (Visual)	4.7 (SFO)	Arnold et al. / 1976
Geomean / Median				8.3 / 10.4