Degradation of fluchloralin in soil under predominating anaerobic conditions

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Degradation offluchloralin in soil underpredominating anaerobicconditionsS.B. Singh a & G. Kulshrestha aa Division of Agricultural Chemicals , IndianAgricultural Research Institute , New Delhi,110012Published online: 21 Nov 2008.

To cite this article: S.B. Singh & G. Kulshrestha (1995) Degradation offluchloralin in soil under predominating anaerobic conditions, Journal ofEnvironmental Science and Health, Part B: Pesticides, Food Contaminants,and Agricultural Wastes, 30:3, 307-319

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J. ENVIRON. SCI. HEALTH, B30(3), 307-319 (1995)

DEGRADATION OF FLUCHLORALIN IN SOIL UNDER PREDOMINATINGANAEROBIC CONDITIONS

Key Words: Herbicide, Fluchloralin, Anaerobicdegradation, Metabolites

S. B. Singh and G. Kulshrestha

Division of Agricultural Chemicals,Indian Agricultural Research Institute,

New Delhi-110012.

ABSTRACT

Degradation of fluchloralin [N-(2-chloroethyl)-

2,6- dinitro-N-propyl-4-(trifluoromethyl)aniline] in

soi l was studied in laboratory under aerobic and

flooded anaerobic conditions. The herbicide degraded

faster in anaerobic than in aerobic soil. The amendment

of flooded anaerobic soil with organic matter further

enhanced the degradation. The major degradation

products ident i f ied were p a r t i a l l y dealkylated

fluchloralin, partially reduced fluchloralin and i ts

cyclized product.

C o n t r i b u t i o n No. 538

307

Copyright © 1995 by Marcel Dekker, Inc.

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308 SINGH AND KULSHRESTHA

INTRODUCTION

Fluchloralin [ N-(2-chloroethyl)-2,6-dinitro-N-p

propyl-4-(trifluoromethyl ) an i l ine , I , Basalin ] , a

substituted dini t roani l ine , is a preemergence , so i l

applied herbicide used for the destruction of various

weeds in wide variety of crops (Rao, 1983) . This

herbicide controls various broad leaf and grassy weeds

(Verma et al., 1978; Rao and Gupta, 1981; Swant and

Jadhav, 1987; Chungi and Ramteke, 1987) commonly

in fe s t i ng r i ce {Oryza sativa), one of the most

important cereal crops in India. In rice field, because

of the standing water, anaerobic conditions normally

prevail . Anaerobic degradation of d in i t roan i l ine

herbicides generally proceeds faster than aerobic

degradation of the same compound (Hellings, 1976;

Probst at al. , 1975) . The present communication

highlights anaerobic degradation of f luchlorol in in

soil .

MATERIALS AND METHODS

Metabolism S t u d i e s

T e c h n i c a l grade f l u c h l o r a l i n was p rov ided by BASF,

I n d i a , New D e l h i . The s o i l used was sandy loam having

60.2% sand, 18.6% c l a y , 21.2% s i l t and 0.35% o r g a n i c

carbon wi th pH of 8 . 2 . Three types of exper iments were

carried out.

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DEGRADATION OF FLUCHLORALIN 309

In the first type, 1 kg sandy loam soil and 20 g

ground straw were placed in a glass bottle (2.50 L) and

1 L of distilled water was added to it to create

flooded conditions. The bottle was closed tight with

cork and kept at room temperature. After four days when

the supernatent water in the bottle became dark brown

in colour indicating the fall in redox potential, the

herbicide fluchloralin (10 mg) was added as a

concentrated ethanol solution (1 ml). The bottle was

again corked and kept in dark at room temperature

(30+1 C] . The experiment was conducted together with

blank soil (without fluchloralin).

In the second experiment soil was not amended

with straw and the soil under flooded conditions was

incubated with and without the herbicide. The third set

of experiment was conducted with aerobic soil. The soil

with and without fluchloralin was maintained with

moisture at field capacity (16%) . The experiments

without herbicide served as control.

All the three experiments were winded up after

seven days. In case of flooded soil the suspension was

centrifuged. The soil was extracted by stirring 1 h

with methanol (1500 ml). The methanol extract was

concentrated to about 25 ml on a rotary evaporator and

was then partitioned between aqueous sodium chloride

and ethyl acetate. The water portion was separately

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extracted with ethyl acetate : hexane (2:8) mixture

(500 ral). This extract was combined with organic layer

of soil extract, washed with water, dried over sodiuri

sulphate and concentrated. The clean up was done by

eluting the concentrate on a column of silica gel with

benzene. Finally, benzene was removed on rotatory

evaporator, the residue dissolved in acetone : hexane

(2:8) and analysed by GLC and TLC.

In case of aerobic studies, the soil was extracted

with IL acetone (500 + 300 + 200 ml) and passed•through

a column of anhydrous sodium sulphate. The extract was

concentrated, diluted with acetone : hexane (2:8) and

analysed by GLC and TLC.

Preparation of Metabolites

2 , 6-Dinitro-H- (propyl ) -4-trif luoroinethylaniline (II )

Fluchloralin (I, Figure 1; 0.5 g) was dissolved in

minimum amount of ethyl alcohol (10 ml) and 20%

potassium hydroxide aqueous solution (3 ml) was added

to it. The contents were refluxed for 6-8 h on a

boiling water bath. The mixture was concentrated to

minimum amount and then diluted with water (10 ml) ,

extracted with ethylacetate (50+50+30 ml), organic

layer washed with water and dried over anhydrous sodium

sulphate. Evaporation of the solvent gave a dark brown

residue which was chromatographed over silica gel.

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C3H7

CIH4C2 C 3 H 7

KOH (aq)/C2H5OH02 N N02

N02

n+m+Ez:

FIGURE 1

Products of fluchloralin degradation in flooded soiland their synthesis

Elution with benzene : hexane (3:7) gave a light brcwr.

thick oil which solidified to give II, m.p., 60 C ,.

literature3 rn.p., 60°C (Leitis and Crosby, 1974!.

2-Amino-N-(2-chloroethyl)-6-nitro-N-propyl-4-trifluoro-

methyl aniline (III)

A solution of fluchloralin (I, 0.5 g) in ethyl

alcohol (25 ir.l) was stirred at room temperature and

treated with 15 ml of 10% aqueous ammonium sulphide

dropwise during a period of 0.5 h. The mixture was

stirred for 2 h and then partitioned between aqueous

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s o d i u m c h l o r i d e and d i c h l o r o i t i e t h a n e . The

dichloromethane l a y e r was washed wi th wa te r , d r i e d over

s o d i u m s u l p h a t e , c o n c e n t r a t e d t o d r y n e s s and t h e

r e s i d u e s u b j e c t e d t o column chromatography over s i l i c a

g e l . E l u t i o n of the column wi th hexane, hexane¡benzene

(7:3) and hexane: benzene (1:1) gave t h r e e f r a c t i o n s in

s u c c e s s i o n .

Concen t ra t ion of hexane f r a c t i o n gave I as yel low

coloured s o l i d (0.04 g) in .p . , 42 C. This was comparable

with f l u c h l o r a l i n on TLC and GLC.

C o n c e n t r a t i o n of h e x a n e : b e n z e n e ( 7 : 3 ) f r a c t i o n

gave IV as a dark brown co loured s o l i d (0 .21 g ) , m . p . ,

68-70 C, Mass s p e c t r u m , e l e c t r o n i m p a c t , m / e , 289

( M + ) ; 1H-MMR (CDCI3) S : 0 . 9 0 ( t , J=7 Hz, 3H,

-CH2CH2Ç_H3) ; 1 .25-1.80 (m, 2H, -CH2Ç_H2CH3 ) ; 2 .85 ( t ,

J=7 Hz, 2H, -CH2CH2CH3); 3 .30 ( s , 4H, -NCH^HgN-) ;

4.25 (bs , 1H, exchangeable wi th D2O, ^NH-) ; 6.73 (d,

J=1.5 Hz, 1H, H-3 Ar; 7.30 (d, J = l . 5 Hz, 1H, H-5 A r ) .

On t h e b a s i s of a b o v e d a t a IV was a s s i g n e d t h e

s t r u c t u r e as 6 - t r i f l u o r o - m e t h y l - 8 - n i t r o - 1 - p r o p y l -

1 , 2 , 3 , 4 - t e t r a h y d r o q u i n o x a l i n e .

C o n c e n t r a t i o n of h e x a n e : b e n z e n e ( 1 : 1 ) f r a c t i o n

gave I I I as a t h i c k brown o i l (O . l lg ) -̂H-NMR (CDCI3) 8

: 1.00 ( t , J=7 Hz, 3H, -CH2CH2CH3); 1 .23-1.92 (m, 2H,

-CH2ÇH2CH3) ; 2 .48-3 .52 (m, 6H, -ÇH2CH2CH3 & -Ç2H_4C1);

3.80 ( s , 2H, exchageable wi th D2O, -NH2); 6.18 and

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6.28 (each d, J=1.50 Hz, each 1H, 2 Ar-H) . On the

basis of ^H-NMR III was assigned the structure as • 2-

amino-N-( 2-chloroethyl )-6-nitro-N-propyl-4-trifluoro

methyl aniline.

Analysis

M.P.s were uncorrected and taken in sulphuric acid

bath. Nuclear Magnetic Resonance (NMR) spectra we're

recorded on a varian EM-360 (60 MHz) instrument using

TMS as an internal standard. Mass spectra was obtained

using a Jeol JMS-D-300 mass spectrometer at 70 ev using

electron impact ionisation with the source at ambient

temperature. GLC was conducted on a Hewlett Packard

Model 5890 II gas Chromatograph fitted with a 6:3Ni

electron capture detector and a megabore column (0.53

mm i.d. x 10 M) packed with HP-1, 2.65 urn film

thickness. The column, injection port and detector,

temperatures were 170, 200 and 300, respectively.

Standard solutions of 0.5 ug/ml each of fluchloralin

(I) , 2 , 6-dinitro-N-propyl-4-trif luoromethylani line

(II), 2-amino- N-(2-chloroethyl)-6-nitro-N-propyl-4~

trifluoromethyl aniline (III) and 6-trifluromethyl-8-

nitro-l-propyl-1,2,3,4-tetrahydroquinoxaline (IV) were

injected into GLC column and the corresponding

retention times recorded were 2.43, 1.68, 3.63 and 5.41

min (Figure 2) respectively. A 3 ul aliquot of cleaned

up soil samples was also injected into GLC. The

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10 min. 10 min. 10 min.

FIGURE 2

Gas c h r o m a t o g r a m s of A: s t a n d a r d s I ( 2 . 4 3 2 ) , I I( 1 . 6 8 0 ) , I I I (3.632) and IV ( 5 . 4 0 8 ) ; B: c o n t r o l f loodeds o i l ; C: f looded s o i l t r e a t e d wi th f l u c h l o r a l i n

residues of fluchlorolin and i t s der ivat ives were

detected by comparing Rts of the samples with

corresponding external standards. The recoveries of I,

I I , III and IV from fortified samples of flooded soil

ranged between 65-70, 65-70, 75-80 and 75-80%

respectively, while from non flooded soil were from 95-

97%. TLC of the standards and flooded soil extract was

developed in benzene using iodine as visualising agent.

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TABLE 1.

Residue of Fluchloralin in Non Sterile Non Flooded,Flooded and Flooded amended with Organic MatterSandyloam Soil.

Incuba-tionperiod

(Days)

07

Non floodedwithout O.M

9.77.5

+ 1.01+ 0.68(25%)

Residue* in ug/g

FloodedWithout O.M

9.5 +1.020.00073 + 0

(99

soil

.0001

.92%)

Floodedwith O.M.

9.4 +1.31Nil(100%)

Figures in parantheses indicated the percentdissipation* Average of three replicatesO.M. - Organic matter

RESULTS AND DISCUSSION

GLC analysis of the extracts of soil samples

showed that the degradation of fluchloralin was faster

in flooded than in non flooded soil. In seven days

99.92% fluchloralin dissipated from flooded soil while

only 25% in non flooded soil. The degradation was

further enhanced and was 100% in the flooded soil

amended with organic matter during the same period

(Table 1). When flooded soil was mixed with organic

matter the degradation of fluchloralin was faster than

before as not even trace of herbicide could be

recovered after four days.

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m

Ts C¡ IV 01 ï

FIGURE 3

Thin layer chromatogram of treated soil (T ) , controlsoil (cg) and standards IV, III, II and I

Various degradation products formed were

identified on the basis of comparison with authentic

samples. GLC (Figure 2) as well as TLC (Figure 3)

showed that in flooded soil the fluchloralin was

transformed into three metabolic products. These were

identified as II, Rt 1.68 min; III, Rt 3.63 min and IV,

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TABLE 2 .

Retardation Factor (Rf) Values of Fluchloralin and itsDerivatives.

Compound Compound TLCnumber name (Rf)

I Fluchloralin 0.86II Dinitropropyl aniline der. 0.13III 6-Aminofluchloralin 0.17IV Quinoxaline derivative 0.31

* Developing solvent: benzene

TABLE 3.

R e t e n t i o n Time of F l u c h l o r a l i n and i t s Th reeDegradation Products on GC f i t t e d with Electron Capture;Detector .

Column Temprature{ C) M2 flow Retention time(Packing) Col/Inj /Det ml/min I I I I I I IV

Megabore 170/200/300 20 2.43 1.68 3.63 5.41(HP-1)

190/210/300 20 1.18 0.92 1.69 2.49

Glass 180/250/275 36 8.50 5.42 12.30 14.80(OV-225),, ' . 220/250/275 36 2.92 2.15 9.36 10.51

Rt 5.41 min. Besides these , two minor products were

also observed on GLC ( Rt, 0.819 and 1.425 min) which

however could not be identified. Retardation factor (Rf

values) of fluchloralin and its derivatives are given

in Table 2. The presence of the degradation products

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II, III and IV was further confirmed by analysing the

soil samples on GLC using columns of different polarity

and different temperature conditions on both the

columns (Table 3). Though product II (21%) and III

(32%) were found to be the major degradation products,

compound IV (2%) was identified as minor product in

soil under flooded anaerobic conditions. However, under

aerobic conditions presence of different metabolites

could not be established in seven days. This could be

attributed to slow degradation of fluchloralin in

aerobic soil.

Formation of products II, III and IV (Figure 1) in

flooded soil indicated two major mechanism of

transformation of fluchloralin in anearobic condition,

partial N- dealkylation and partial reduction followed

by cyclisation. The presence of organic matter enhanced

these reactions. In aerobic soil fluchloralin persisted

for a longer period and only 25% was dissipated in

seven days.

ACKNOWLEDGEMENTS

The authors thank Dr. N.K.Roy, Head, Division of

Agricultural Chemicals, New Delhi for providing

facilities.

REFERENCES

Chungi, M. D. and Ramteke, J. R., Annals Agricul. Res.8, 166-168 (1987).

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Helling, C.S., J. Environ. Qual., 5 , 1-12. (1976).

Leitis, E. and Crosby, D. G., J. Agric. Food Chem., 22,842-848 (1974).

Probst, G. W., Golab, T. and Wright, W.L. "Herbicides:Chemistry, Degradation and Mode of action", Kearney, P.C. and Kaufman, D. D., eds., Marcel Dekker, Inc., NewYork (1975) pp 453-500.

Rao, K.N., and Gupta, K.M. Weed Abstr., 25119 (1984).

Rao, V.S. "Principles of Weed Science", Oxford & IBHPublishing Co., New Delhi, India (1983) pp 384-392.

Verma, O.P.S., Tyagi, R.C. and Katyal, S.K.,Pesticides, 12, 21-22 (1978).

Swant, A.C. and Jadhav, S.N., Indian Weed Sci., 17, 35-39 (1987).

Received: February 22, 1994

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Documents

Degradation of fluchloralin in soil under predominating anaerobic conditions