Pest Management Science Pest Manag Sci 56:1065±1072 (2000)

The soil degradation of the herbicide florasulamRoy Jackson,* Dipankar Ghosh and Glen PatersonDow AgroSciences, Letcombe Regis, Wantage, Oxfordshire OX12 9JT, UK

(Rec

* CoE-ma

# 2

Abstract: The route and rate of degradation of ¯orasulam, a low-rate triazolopyrimidine sulfonanilide

herbicide, was investigated in six soil types under aerobic conditions at 20 or 25°C. Degradation

products were isolated and identi®ed by mass spectroscopy. Florasulam was rapidly degraded by

microbial action with an average half-life of 2.4 days (range 0.7 to 4.5 days). The ®rst step in the

degradation pathway involved conversion of the methoxy group on the triazolopyrimidine ring to a

hydroxy group to form N-(2,6-di¯uorophenyl)-8-¯uoro-5-hydroxy[1,2,4]triazolo[1,5-c]pyrimidine-2-

sulfonamide. This metabolite degraded, with a half-life of 10 to 61 days, via partial breakdown of the

triazolopyrimidine ring to form N-(2,6-di¯uorophenyl)-5-aminosulfonyl-1H-1,2,4-triazole-3-car-

boxylic acid. This was followed by cleavage of the sulfonamide bridge to form 5-(aminosulfonyl)-

1H-1,2,4-triazole-3-carboxylic acid. Other degradation processes involved decarboxylation of the

carboxylic acid metabolites and mineralisation to form carbon dioxide and non-extractable residues.

# 2000 Society of Chemical Industry

Keywords: ¯orasulam; DE-570; triazolopyrimidine sulfonanilide; soil; degradation; environmental fate

Figure 1. Structure of florasulam showing (a) 14C-phenyl and (b) 14C-TPlabel position.

1 INTRODUCTIONFlorasulam (Dow AgroSciences code DE-570; N-

(2,6-di¯uorophenyl)-8-¯uoro-5-methoxy[1,2,4]tri-

azolo[1,5-c]pyrimidine-2-sulfonamide; Fig 1) is a tri-

azolopyrimidine sulfonanilide herbicide for the control

of broad-leaf weeds in cereals.1±3 The mode of action

is through inhibition of the enzyme acetolactate

synthase (ALS) by foliar or root uptake following

post-emergence application. Florasulam is particularly

active on Galium aparine L, Stellaria media (L) Vill,

Matricaria spp and various cruciferae at very low rates,

typically 2.5 to 7.5ghaÿ1. This application rate will

result in an extremely low environmental loading and

very low residues in crops and soil. However, because

of the high herbicidal activity of ¯orasulam on broad-

leaf plants, it is important to assess any potential for

damage to following crops and other non-target plants.

A fundamental part of this assessment is to determine

how rapidly ¯orasulam will degrade in soil. It is also

important to identify the major degradation products

so that their potential impact on the environment can

be assessed.

In this study, the degradation of 14C-labelled ¯ora-

sulam in different agricultural soil types under aerobic

conditions was investigated and the major degradation

products were identi®ed by high performance liquid

chromatography and mass spectroscopy. The very low

application rate of ¯orasulam presented a major

challenge with respect to the quanti®cation and identi-

®cation of these metabolites and the use of a 1:1 ratio

of 12C:14C facilitated identi®cation by mass spectro-

scopy and radio-detection.

eived 19 May 2000; revised version received 20 July 2000; accepted

rrespondence to: Roy Jackson, Dow AgroSciences, Letcombe Regis,il: jacksonr@dow.com

000 Society of Chemical Industry. Pest Manag Sci 1526±498X/2

2 MATERIALS AND METHODS2.1 SoilsSix soils (four European and two North American)

with a range of physico-chemical properties were used

to investigate the degradation of ¯orasulam (Table 1).

The soils were collected from the top 20cm of the ®eld

and stored under aerobic conditions in the dark at

approximately 4°C prior to use, to preserve microbial

activity. The soils were sieved through a 2-mm screen

and were used within three months of collection. A

sample of one of the soils (Speyer 2.2 loamy sand) was

sterilised by gamma irradiation in order to investigate

the relative importance of biotic and abiotic degrada-

tion.

2.2 Test material and reference standardsTwo batches of [14C]¯orasulam radiolabelled in

different positions were supplied by the Speciality

Synthesis group of Dow AgroSciences, Indianapolis,

7 August 2000)

Wantage, Oxfordshire OX12 9JT, UK

000/$30.00 1065

Table 1. Soil characterisation data

Soil Sand a (%) Silt a (%) Clay a (%) Organic C (%) CEC (mEq 100gÿ1) pH Moisture b (%)

Speyer 2.2 loamy sand (Germany) 76.3 15.5 8.2 3.9 8.7 7.3 25.6

Marcham sandy clay loam (UK) 48.2 23.0 28.8 2.0 9.0 7.7 21.8

Kenslow humus silt loam (UK) 13.5 62.4 24.1 6.8 16.0 5.6 38.0

Andover silt loam (UK) 18.5 59.2 22.3 3.1 12.0 7.6 25.7

Hanford sandy loam (USA) 60.0 31.2 8.8 1.0 5.2 7.4 8.5

Catlin silty clay loam (USA) 10.8 62.0 27.2 2.2 15.3 7.0 25.4

a Sand =2mm±63mm (European soils); 2mm±50mm (US soils). Silt=63±2mm (European soils); 50±2mm (US soils). Clay=< 2mm.b Expressed as % moisture per dry weight of soil.

R Jackson, D Ghosh, G Paterson

USA. One batch was uniformly labelled in the phenyl

(Ph) ring with a speci®c activity of 2020MBqmmoleÿ1

and the other was labelled at the 9-position of the

triazolopyrimidine (TP) ring with a speci®c activity of

2065MBqmmoleÿ1 (Fig 1). The radiochemical purity

of both test materials was > 97%. Non-radiolabelled

¯orasulam and the metabolite reference standards

N-(2,6-di¯uorophenyl)-8-¯uoro-5-hydroxy[1,2,4]tri-

azolo[1,5-c]pyrimidine-2-sulfonamide, N-(2,6-di-

¯uorophenyl)-5-aminosulfonyl-1H-1,2,4-triazole-3-

carboxylic acid (DFP-ASTCA), N-(2,6-di¯uorophe-

nyl)-1H-1,2,4-triazole-3-sulfonamide (DFP-TSA), 5-

(aminosulfonyl)-1H-1,2,4-triazole-3-carboxylic acid

(ASTCA) and 1H-1,2,4-triazole-3-sulfonamide

(TSA) were also supplied by the Speciality Synthesis

group of Dow AgroSciences.

2.3 Experimental procedureDegradation of ¯orasulam was investigated in glass

biometer ¯asks consisting of two joined conical ¯asks

similar to the system described by Bartha and Pramer.4

Samples of soil (50g dry weight equivalent) were

weighed into one side of the biometer ¯asks and the

moisture content was adjusted to either 40% of the

moisture holding capacity (European soils) or 75% of

the moisture holding capacity at an applied suction

pressure of 1/3 bar (North American soils). In general,

a moisture content of 75% at 1/3 bar is drier than 40%

moisture holding capacity. The [14C]¯orasulam was

dissolved in distilled water and applied to soil at a

nominal application rate of 0.015mg kgÿ1 for the

European soils and 0.024mg kgÿ1 for the North

American soils. The predicted soil concentration

following application at 7.5ghaÿ1 (assuming incor-

poration to 5cm and a soil density of 1.5gcmÿ3) was

0.01mg kgÿ1 but the slightly higher rate was used to

aid quantitation. The ¯asks were shaken to distribute

the test substance in the soil. Sodium hydroxide

solution (0.1M, 100ml) was added to the side-arm of

each biometer ¯ask to trap evolved carbon dioxide. To

maintain aerobic conditions, the ¯asks were connected

to a low-pressure oxygen supply so that the oxygen

removed as a result of soil respiration was replaced.

The soils were maintained in the dark at either

20(�2)°C (European soils) or 25(�2)°C (North

American soils).

Biometer ¯asks were analysed at various time points

up to 100 days for the European soils and up to 1 year

1066

for the North American soils. The soils were initially

extracted by shaking for 10min with aqueous calcium

chloride solution (0.01M; 100ml). After centrifugation

and removal of the aqueous supernatant, the soil was

further extracted with acetone�water�acetic acid

(90�9�1 by volume; 3�100ml) by shaking for 1h.

From day 7 onwards, a more vigorous extraction pro-

cedure was required to extract 14C-residues from soil.

Therefore, soils were further extracted with aceto-

nitrile�water�phosphoric acid (80�19�1 by

volume; 2�100ml) by shaking for 1h. Soil extracts

were analysed by liquid scintillation counting (LSC) to

determine the total radioactivity in each extract.

Soil extracts were concentrated using solid phase

extraction (SPE) procedures. Each aqueous calcium

chloride extract was acidi®ed to pH 2 and passed

through a C18 SPE column. The column was eluted

with methanol, which was evaporated to a small

volume (0.5±1.0ml) under nitrogen at 40°C. The

organic soil extracts were evaporated to a small volume

(10±20ml) to remove the organic solvent and sub-

jected to a similar SPE procedure. The organic and

aqueous fractions after SPE were analysed by LSC to

determine recoveries. At later time points, a signi®cant

proportion of radioactivity remained in the aqueous

phase from solid phase extraction of extracts of soil

treated with the 14C-TP label only. These aqueous

fractions were freeze-dried and the residue was re-

dissolved in 1±2ml of water�acetonitrile (95�5)

containing 10ml litreÿ1 acetic acid.

Soil residues after extraction were air-dried, mixed

thoroughly and triplicate samples (c 0.4±0.5g) were

combusted in oxygen using a Harvey OX500 biologi-

cal sample oxidiser.

2.4 Measurement of radioactivityTriplicate samples of soil extracts (0.5 or 1ml),

sodium hydroxide trapping solutions (2ml) and other

liquid samples generated during the study (eg con-

centrated extracts, HPLC eluate) were mixed with

Ultima-Gold2 XR (Packard) scintillation cocktail.

The [14C]carbon dioxide resulting from combustion

of soils was absorbed in a 1�1 mixture of Carbo-

Sorb2 (Packard) carbon dioxide absorbing solution

and Perma¯uor2 (Packard) scintillation cocktail. All

samples were counted for 5min using a Beckman LS

3801 liquid scintillation counter with external quench

correction.

Pest Manag Sci 56:1065±1072 (2000)

Soil degradation of ¯orasulam

2.5 Chromatographic analysisAll soil extracts were analysed by high performance

liquid chromatography (HPLC). Instrumentation

consisted of a Varian 9010 pump, a Waters auto

sampler (Model 712 or 717), a Varian 9050 UV

detector (set at 260nm) and either a Berthold LB506

or a Packard Radiomatic 500TR radioactivity detector

with associated data collection software. Two different

separation systems were used. System 1 used an

octadecylsilyl (ODS) bonded silica column (Spheri-

sorb 5 ODS2, 25cm�4.6mm, Hichrom). The mobile

phase consisted of acetic acid�water (2�98 by

volume; solvent A) and acetic acid�acetonitrile

(2�98 by volume; solvent B) with the following

elution gradient: T0 (min)=100:0 (A:B), T7=100:0,

T25=50:50, T35=0:100, T45=0:100. The ¯ow rate

was 1ml minÿ1. System 2 used a porous graphitic

carbon (PGC) column (Hypercarb 7mm, 5cm�4.6mm, Shandon). The mobile phase consisted of

tri¯uoroacetic acid�water (1�99 by volume; solvent

A) and tri¯uoroacetic acid�acetonitrile (1�99 by

volume; solvent B) with the following elution gradient:

T0 (min)=95:5 (A:B), T30=30:70, T35=0:100,

T45=0:100. The ¯ow rate was 1ml minÿ1.

2.6 Isolation of metabolitesTo generate suf®cient amounts of metabolites for

identi®cation, samples of soil (50g) were treated with

[14C]¯orasulam at 1.5mg kgÿ1 (ie 100 times the

application rate used for the main study). Samples of

soil were also treated with a 1�1 mixture of radio-

labelled and non-radiolabelled ¯orasulam. This dilu-

tion of the radiolabel was designed to facilitate analysis

by mass spectroscopy since it gave a characteristic 1:1

(12C:14C) doublet in the mass spectra which allowed

sample-related ions to be more easily identi®ed. The

soils were maintained at 20°C under aerobic condi-

tions for 62 days. The soils were extracted with

acetone�water�concentrated hydrochloric acid

(90�9�1 by volume; 100ml). The extracts were

evaporated to a small volume and separated into

organic and aqueous fractions by SPE. The fractions

were analysed by HPLC and fractions of column

eluate containing the various metabolites were col-

lected.

2.7 Mass spectroscopic analysisThe samples were analysed by electrospray liquid

chromatography mass spectrometry (ESI LC-MS),

with of¯ine 14C radioactive detection. The mass

spectrometer was a Finnigan-MAT TSQ700 equipped

with Finnigan-MAT electrospray (ESI) interface. The

mass spectrometer was linked to a HP1050 LC system

and a Berthold 14C radioactive monitor (RAM). The

LC system consisted of a Hewlett-Packard pump,

auto-sampler and de-gasser connected to a Prodigy

ODS3 column (25cm�4.6mm, Phenomenex Ltd).

The mobile phase contained acetic acid�water

(1�99 by volume; solvent A) and acetic acid�acetonitrile (1�99 by volume; solvent B) with two

Pest Manag Sci 56:1065±1072 (2000)

different elution gradients depending on which metab-

olites were being analysed. For the DFP-ASTCA and

DFP-TSA metabolites, the following gradient was

used: T0 (min)=95:5 (A:B), T30=30:70, T35=

0:100, T45=0:100. For the more polar ASTCA and

TSA metabolites, the gradient was: T0 (min)=100:0

(A:B), T5=100:0, T20=50:50, T40=0:100.

All analyses were carried out by ESI LC-MS in the

negative ion mode using the following mass spectro-

metry conditions: capillary temperature, 250°C; spray

voltage, 4.5±5.5kV; spray current, Variable (c 30±

70mA); scan range, 150±700amu in 1s.

2.8 Degradation kinetics analysisDegradation of ¯orasulam was described by ®rst-order

kinetics according to the following equation:

�C�t � �C�0 � eÿkt

where [C]0 is the initial concentration of ¯orasulam in

soil, [C]t is the concentration at time t and k is the ®rst-

order rate constant. A plot of ln [C] against time was

made and the best-®t line was determined by linear

regression. The slope of this line is equal toÿk and the

®rst-order degradation half-life (t1/2) was calculated

from ln (2)/k.

3 RESULTS3.1 Recovery and distribution of radioactivityThe distribution of radioactivity in one European soil

(Speyer 2.2 loamy sand) and one North American soil

(Catlin silty clay loam) is presented in Tables 2 and 3,

respectively. Total recoveries were generally greater

than 90% of applied radioactivity (AR) indicating that

there was no signi®cant loss of 14C during the

experiment. Total extractable 14C-residues decreased

with time, particularly in soils treated with the 14C-

phenyl label. Extractability decreased to 52% AR after

1 year in Catlin soil treated with TP label compared

with 11% in the same soil treated with the Ph label.

Mineralisation to carbon dioxide was signi®cantly

higher for the Ph label than for the TP label (47%

and 23% AR, respectively, after 1 year in Catlin soil).

3.2 Degradation of florasulam and identification ofthe primary metaboliteThe degradation of ¯orasulam is shown graphically in

Fig 2 and calculated half-lives are presented in Table

4. Degradation was rapid in all six soils and showed a

good ®t with pseudo-®rst-order kinetics (R2>0.95)

with half-lives in the range 0.7 to 4.5 days. Degrada-

tion in sterile soil was much slower (half-life=116

days) indicating that degradation was predominantly a

microbial process.

The primary degradation product was N-(2,6-di-

¯uorophenyl)-8-¯uoro-5-hydroxy[1,2,4]triazolo[1,5-

c]pyrimidine-2-sulfonamide (5-OH-¯orasulam)

formed by conversion of the methoxy group to a

hydroxy group. The structure of this metabolite

1067

Table 2. Distribution of radioactivity in the Speyer 2.2 European soil treated with [14C]florasulam

Label a Time (days) Total extract Florasulam 5-OH DFP-ASTCA ASTCA Other b 14CO2 Soil residue Total recovery c

TP 0 96.4 91.8 3.8 nd nd 0.8 ns 2.1 98.5

1 94.1 39.6 49.9 1.1 0.8 2.7 nd 4.0 98.1

3 88.1 22.6 55.8 4.1 nd 5.6 nd 7.6 95.7

7 84.3 4.6 61.1 9.5 3.8 5.3 <0.1 13.7 98.0

14 73.4 1.2 41.0 15.3 7.0 9.0 0.1 23.1 96.6

28 55.5 0.6 21.9 17.5 8.0 7.5 0.6 39.6 95.7

59 49.4 nd 11.8 18.1 15.7 3.9 1.1 41.6 92.2

100 39.4 nd 3.5 8.8 26.1 0.9 4.9 48.3 92.5

Ph 0 96.7 96.2 nd 0.2 nd 0.3 ns 1.8 98.5

1 95.2 42.2 52.7 nd nd 0.3 0.1 3.4 98.7

3 87.3 19.6 65.3 1.8 nd 0.5 0.4 9.6 97.3

7 78.9 4.7 64.2 8.3 nd 1.7 1.8 15.8 96.5

14 65.4 2.6 44.5 16.6 nd 1.8 3.1 26.5 95.1

28 47.4 0.3 23.7 20.5 nd 3.0 7.1 40.4 95.0

59 30.8 0.2 9.2 18.6 nd 2.9 11.0 48.4 90.2

100 20.8 nd 5.5 15.2 nd 0.2 13.6 57.1 91.5

Sterile TP 1 96.7 79.3 12.4 0.3 nd 4.7 nd 3.2 99.9

8 94.5 77.3 7.9 nd nd 9.3 nd 5.8 100.2

28 88.8 66.5 8.4 1.2 0.7 12.1 <0.1 7.1 96.0

106 77.9 44.1 17.6 2.4 1.1 12.8 0.1 17.4 95.4

nd=not detected, ns=no sample.a TP=triazolopyrimidine 14C label, Ph=phenyl 14C label.b Other consists of several minor metabolites�aqueous-soluble radioactivity not analysed by HPLC.c Total recovery is sum of extract, residue and 14CO2.

R Jackson, D Ghosh, G Paterson

(molecular mass 345amu) was con®rmed by mass

spectroscopic and chromatographic comparison with a

synthesised reference standard. The formation and

Table 3. Distribution of radioactivity in Catlin US soil treated with [14C]florasulam

Label a Time (days) Total extract Florasulam 5-OH DFP-AST

TP 0 97.6 90.9 6.1 nd

1 95.4 66.0 28.8 nd

3 96.0 20.9 65.8 4.9

7 91.4 6.1 70.4 6.3

14 83.5 2.2 60.5 10.8

28 68.5 1.9 36.5 8.2

56 60.1 1.5 22.2 3.9

91 45.9 0.7 9.6 1.2

182 50.6 nd 5.8 nd

273 48.1 nd 2.7 nd

364 52.3 nd nd nd

Ph 0 96.1 91.8 4.3 nd

1 93.0 67.5 25.5 nd

3 95.4 30.0 62.3 2.2

7 81.2 7.2 67.6 5.5

14 70.4 1.8 59.2 8.8

28 51.2 nd 34.5 4.6

56 33.2 nd 28.6 3.5

91 18.8 nd 17.0 0.1

182 12.9 nd 9.9 nd

273 11.1 nd 10.1 nd

364 10.6 nd 10.3 nd

nd=not detected, ns=no sample.a TP=triazolopyrimidine 14C label, Ph=phenyl 14C label.b Other consists of several minor metabolites�aqueous-soluble radioactivity not ac Total recovery is sum of extract, residue and 14CO2.

1068

decline of this metabolite is shown in Fig 3. Maximum

levels of the 5-OH metabolite (41±72% AR) were

observed between 3 and 7 days after application. First-

CA ASTCA Other b 14CO2 Soil residue Total recovery c

nd 0.6 ns 0.4 98.0

nd 0.7 <0.1 2.9 98.3

1.6 2.7 <0.1 2.7 98.7

1.4 6.4 0.2 4.2 95.8

5.0 5.0 1.7 7.8 93.0

17.6 4.2 5.8 13.7 88.0

23.4 9.1 13.7 17.3 91.1

25.1 9.1 16.1 17.3 79.3

24.7 8.6 17.5 21.1 89.2

38.4 3.3 21.2 23.1 92.4

43.3 7.9 22.9 24.2 99.4

nd nd ns 0.3 96.4

nd nd 0.1 3.8 96.9

nd 0.5 0.7 3.6 99.7

nd 0.9 3.1 7.6 91.9

nd 0.6 7.4 14.4 92.2

nd 0.9 16.0 23.8 91.0

nd 0.6 22.9 32.9 89.0

nd 0.9 31.4 34.3 84.5

nd 1.1 40.0 31.8 84.7

nd 0.6 44.0 31.6 86.7

nd nd 47.2 29.9 87.7

nalysed by HPLC.

Pest Manag Sci 56:1065±1072 (2000)

Figure 3. Formation and decline of the 5-hydroxy metabolite in soil.Figure 2. Decline of florasulam in six soil types.

Soil degradation of ¯orasulam

order half-lives for the metabolite, estimated by linear

regression analysis of the decline curve, are presented

in Table 3. Degradation was relatively rapid, with half-

lives of 10±31 days in the four European soils. Longer

half-lives (56±62 days) were observed in the USA soils,

which may have been due to the drier condition of

these soils.

3.3 Identification and quantification of secondaryand tertiary metabolitesA second metabolite, more polar than both ¯orasulam

and the 5-OH metabolite, was observed with both

labelled forms of [14C]¯orasulam at maximum levels

of 4±22% AR. The negative electrospray mass

spectrum of this metabolite (Fig 4a) showed the

molecular ion [MÐH]ÿ as a 1:1 doublet of m/z 303

(12C) and 305 (14C), indicating a molecular mass of

304amu. The presence of the 12C:14C doublet helped

to identify this ion as a metabolite of ¯orasulam and

the presence of another doublet ion at m/z 259 and 261

corresponded with a loss of 44amu (possibly CO2),

suggesting that the metabolite was a carboxylic acid.

The metabolite was postulated to be N-(2,6-di¯uoro-

phenyl)-5-aminosulfonyl-1H-1,2,4-triazole-3-car-

boxylic acid (DFP-ASTCA). A reference standard of

DFP-ASTCA was synthesised and the soil metabolite

was shown to co-chromatograph with the standard by

HPLC. The mass spectrum of the metabolite (Fig 4a)

matched that of the reference standard (Fig 4b),

con®rming that the postulated structure was correct.

A third metabolite, which was detected only with the

Table 4. First-order soil degradation half-lives (t1/2) for florasulam and its

Soil

Florasulam

t1/2 (days) R2 Data ran

Speyer 2.2 loamy sand 1.7 0.9787 0

Marcham sandy clay loam 4.5 0.9776 0

Kenslow humus silt loam 0.7 0.9983 0

Andover silt loam 1.5 0.9958 0

Hanford sandy loam 2.5 0.9543 0

Catlin silty clay loam 3.3 0.9587 0

Pest Manag Sci 56:1065±1072 (2000)

14C-TP label, accounted for >10% AR in ®ve of the

six soils and was the major metabolite at later time

points. The negative electrospray mass spectrum of

this metabolite showed a molecular ion [MÐH]ÿ of

m/z 191 (12C) and 193 (14C) indicating a molecular

mass of 192amu. This metabolite was postulated to

be 5-(aminosulfonyl)-1H-1,2,4-triazole-3-carboxylic

acid (ASTCA) which is formed by removing the

phenyl ring of DFP-ASTCA. A reference standard of

ASTCA was synthesised and a good match between

the HPLC retention times and mass spectra of the

metabolite and standard con®rmed that the postulated

structure was correct.

Two other metabolites were identi®ed as N-

(2,6-di¯uorophenyl)-1H-1,2,4-triazole-3-sulfonamide

(DFP-TSA) and 1H-1,2,4-triazole-3-sulfonamide

(TSA) which are the decarboxylated forms of DFP-

ASTCA and ASTCA, respectively. These metabolites

were generally seen at low levels and there was

evidence that they were formed by decarboxylation

of DFP-ASTCA and ASTCA during the extraction

procedure. An additional experiment to determine the

stability of these two carboxylic acid metabolites in the

acetonitrile/phosphoric acid extraction solvent showed

that approximately 20% of the initial material de-

graded to the decarboxylated products after one day at

ambient temperature.

The route of degradation of ¯orasulam in soil is

shown in Fig 5. Although cleavage of the sulfonamide

bridge to form ASTCA is a major step in the metabolic

pathway, no corresponding metabolites were formed

5-hydroxy metabolite

5-Hydroxy metabolite

ge (days) t1/2 (days) R2 Data range (days)

±7 25 0.9600 7±100

±14 29 0.8986 7±100

±3 31 0.8962 3±100

±7 10 0.9920 7±59

±14 56 0.9716 14±182

±14 62 0.9156 7±273

1069

Figure 4. Negative ion ESI mass spectrumof (a) soil metabolite showing characteristic12C: 14C doublet for [M—H]ÿ ion at m/z 303and 305 and (b) reference standard ofDFP-ASTCA showing [M—H]ÿ ion at m/z303 (12C).

R Jackson, D Ghosh, G Paterson

from the phenyl half of the molecule. This can be

explained by the higher levels of [14C]carbon dioxide

and non-extractable residues observed for the Ph label,

which indicate that the phenyl ring is rapidly broken

down to these terminal degradation products.

3.4 Extractability of florasulam and its metabolitesA comparison of the distribution of radioactivity in the

aqueous and organic soil extracts gives useful informa-

tion about the potential mobility of ¯orasulam and its

metabolites in the soil environment and their avail-

ability for uptake by plants. This distribution between

the aqueous and organic extracts is shown in Fig 6. It

can be seen that ¯orasulam and the 5-OH metabolite

were extracted mainly with the aqueous extract. The

DFP-ASTCA metabolite was divided more evenly

between the aqueous and organic extracts and almost

all of the ASTCA metabolite was extracted with the

organic solvent. These results suggest that ¯orasulam

and the primary 5-OH metabolite are readily desorbed

from soil, but the metabolites which are likely to be

present at later times are more strongly adsorbed and

will be less mobile and less bioavailable.

1070

3.5 Concentration of florasulam and itsmetabolites in soilBecause of its low application rate, the actual

concentration of ¯orasulam in soil will be low. For

example, the predicted environmental concentration

of ¯orasulam in soil following application at 7.5ghaÿ1

is 0.01mg kgÿ1, assuming a soil density of 1.5gcmÿ3,

incorporation to a depth of 5cm and no crop inter-

ception. This concentration would be expected to

decrease rapidly based on the degradation rates

observed in this study. The predicted maximum levels

of metabolites based on the observed maximum

concentrations in this study are 0.007mg kgÿ1 for

the 5-OH metabolite and 0.002mg kgÿ1 for DFP-

ASTCA and ASTCA.

4 DISCUSSION AND CONCLUSIONSThe degradation of ¯orasulam was investigated in a

wide range of soils under aerobic conditions and the

major metabolites were identi®ed. The use of a 1:1

ratio of 12C:14C facilitated the identi®cation of low-

level metabolites by mass spectroscopy. Florasulam is

Pest Manag Sci 56:1065±1072 (2000)

Figure 5. Route of degradation offlorasulam in soil.

Soil degradation of ¯orasulam

rapidly degraded by microbial processes with an

average half-life of 2.4 days (range 0.7±4.5 days) at

20±25°C. The primary metabolite, 5-OH-¯orasulam,

is also readily degraded with an estimated half-life of

10±62 days. Four other metabolites were identi®ed

resulting from partial breakdown of the triazolopyr-

imidine ring and cleavage of the sulfonamide bridge.

The 5-OH metabolite is known to be at least 4000

times less active on the target enzyme (ALS) than

Figure 6. Distribution of florasulam andmetabolites between aqueous andorganic extracts.

Pest Manag Sci 56:1065±1072 (2000)

¯orasulam (Ehr RJ, Schmitzer PR and Gray JA, pers

comm, 1997) and has been shown to have no pre-

emergence soil activity at up to seven times the

anticipated use rate (Ehr RJ and Alexander AL, pers

comm, 1997). In addition, the secondary metabolites

have been shown to have little or no ALS activity (Ehr

RJ and Alexander AL, pers comm, 1997).

Extraction with aqueous and organic solvents

showed that ¯orasulam and the 5-OH metabolite were

1071

R Jackson, D Ghosh, G Paterson

readily desorbed in aqueous solution. The metabolites

formed at later time points were more strongly

adsorbed to soil and should be less mobile and less

bioavailable than the parent herbicide.

In conclusion, the low application rate of ¯orasulam

(2.5±7.5ghaÿ1) together with the rapid degradation

(half-life=2.4 days) to form non-phytotoxic metab-

olites should lead to very low residues in soil.

ACKNOWLEDGEMENTSThe authors would like to acknowledge the assistance

of Len McKendry and Frank White of Dow Agro-

Sciences in the synthesis of metabolite reference

standards.

1072

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Farnham, Surrey, UK, pp 73±80 (1999).

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