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LIBRARY

Michlgan State
Unlversity

 

 

 

 

This is to certify that the
dissertation entitled
EFFICACY OF ANNUAL GRASS CONTROL BY METOLACHLOR AS
INFLUENCED BY FLUMETSULAM, HALOSULFURON AND
CHLORIMURON-ETHYL

presented by
Karen M. Novosel

has been accepted towards fulfillment
of the requirements for

Ph.D. degreein Crop rind Soil Sciences

 

Major professor

Date 7/ 14% 9 /99fi7
(l /

MS U is an Affirmative Action/Equal Opportunity Institution 0-12771

 

PLACE N RETURN BOX to remove thle checkout from your record.
TO AVOID FINES return on or bdore dete due.

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MSU to An Afflrmettve Action/Equal Opportuntty lnetltwon
Wm!

EFFICACY OF ANNUAL GRASS CONTROL BY METOLACHLOR AS
INFLUENCED BY FLUMETSULAM, HALOSULFURON AND
CHLORIMURON-ETHYL
By

Karen M. Novosel

A DISSERTATION
Submitted to
Michigan State University

in partial fulfillment of the requirements
for the degree of

DOCTOR OF PHILOSOPHY

Department of Crop and Soil Sciences

1997

ABSTRACT
EFFICACY OF ANNUAL GRASS CONTROL BY METOLACHLOR AS
INFLUENCED BY FLUMETSULAM, HALOSULFURON AND
CHLORIMURON-ETHYL
By

Karen M. Novosel

Research was conducted to determine the effect on annual grass control by
metolachlor in the presence of ALS-inhibiting herbicides. In greenhouse studies,
metolachlor at 0.14, 0.28, and 0.56 kg ai ha" was applied alone and in combination with
flumetsulam at 0.0073 and 0.015 kg ai ha" and halosulfuron at 0.011 and 0.021 kg ai ha“.
Combining flumetsulam and halosulfuron with metolachlor did not increase grass control
beyond that observed from metolachl'or alone. Grass control from 0.14 kg ha'1 of
metolachlor was reduced when applied with 0.021 and 0.011 kg ha" of halosulfuron or
0.015 kg ha" of flumetsulam. These interactions were determined to be antagonistic
according to Colby’s multiplicative interactive model. In field studies conducted for 3 yr
in corn and soybean, grass control did not consistently increase or decrease when
flumetsulam, halosulfuron or chlorimuron was applied with metolachlor compared to
metolachlor alone. In greenhouse studies to determine the activity of these ALS inhibitors
on giant foxtail, metolachlor at 0.28 kg ha'1 provided 97% control of giant foxtail 28 days
after treatment. Chlorimuron, halosulfuron plus furilazole, flumetsulam and halosulfuron
provided 63, 60, 44 and 31% control of giant foxtail, respectively. In laboratory

experiments, incidences of antagonism occurred only when an ALS inhibiting herbicide

was applied 24 hours prior to metolachlor. The radicle of giant foxtail emerged in 3 d

while the shoots emerged between 5 and 14 d. Metolachlor and chlorimuron were
predominantly absorbed by giant foxtail shoots while flumetsulam and halosulfuron were
root absorbed. The order of soil mobility was flumetsulam> chlorimuron> halosulfuron
with furilazole=halosulfuron> metolachlor. Seeds pretreated with an ALS inhibitor
absorbed 2 to 5 times more l“C-metolachlor than seeds pretreated with metolachlor,
pretreated with metolachlor plus an ALS inhibitor or not pretreated (metolachlor only).
Exposure to an ALS-inhibiting herbicide 24 h prior to metolachlor caused more rapid and
greater overall metabolism 8 h after treatment. These data indicate that increased
metabolism of metolachlor as a result of exposure to an ALS-inhibiting herbicide could

result in antagonism under field conditions.

ACKNOWLEDGMENTS

I would like to thank Dr. Karen Renner, my major advisor, for all her help and
support throughout my graduate school experience. I appreciate all that you have done fo r
me. Dr. Donald Penner, Dr. James Kells and Dr. Matthew Zabik also provided excellent
guidance as members of my committee by giving their insight and advice which was
immeasurably helpful. Gary Powell made my field work feasible and enjoyable. I would
like to thank Eric Spandl for his help with the initial greenhouse study and for all the
statistics and computer help he gave me. The legacies of Ghandi, J. Boyd, the Hartman
have inspired me to try to attain their immortality. Also, the help and support of my fellow
graduate Students Aaron Hager, Rick Schmenk, Julie Lich, Bob Starke, Jason Fausey,
Kelly Nelson, Corey Ransom, Brent Tharp, Christy Sprague, Matt Rinella, Paulie Knoerr ,
John Burk and Terry Wright made the completion of this dissertation possible. The student
workers who have made my work load that much more bearable deserve acknowledgment
here as well. This great crew of people includes Mike Particka, Angie Eichorn, Kyle

Fiebig and Jim Sherman.

iv

TABLE OF CONTENTS

List of Tables ......................................................................... vii
List of Figures ......................................................................... ix
Introduction ............................................................................. 1

Chapter 1 . Literature Review

Introduction ....................................................................... 2
Metolachlor ........................................................................ 3
Flumetsulam ....................................................................... 8
Chlorimuron-ethyl ............................................................... 10
Halosulfuron and furilazole .................................................... 13
Interactions ........................................................................ 14
Herbicide interactions ............................................................ 15
Literature Cited ................................................................... 17

Chapter 2. Metolachlor Efficacy as Influenced by Three Acetolactate Synthase
Inhibiting Herbicides Under Field and Greenhouse Conditions

Abstract ............................................................................ 21
Introduction ....................................................................... 22
Materials and Methods
Greenhouse Study ....................................................... 23
Field studies .............................................................. 24
Results and Discussion
Greenhouse Results .................................................... 26
Field Results 1994 ...................................................... 26
Field Results 1995 ...................................................... 28
Field Results 1996 ...................................................... 29
Literature Cited ........................................................ 32

Chapter 3. Physiological Basis for the Antagonistic Response of Giant Foxtail
(Setaria faben' L.) to Metolachlor in the presence of Flumetsulam,
Halosulfuron and Chlorimuron-ethyl.

Abstract ........................................................................... 43
Introduction ...................................................................... 44

Materials and methods

Greenhouse Study ....................................................... 46
Pregermination ........................................................... 47
Timing study .............................................................. 47
Emergence study ......................................................... 48
Site of Uptake study ..................................................... 49
Soil mobility studies ..................................................... 50
Absorption study ......................................................... 51
Metabolism study ........................................................ 52
Results and Discussion
Greenhouse Results ...................................................... 53
Timing Results ........................................................... 54
Emergence Results ....................................................... 56
Site of Uptake Results ................................................... 56
Soil mobility Results ..................................................... 57
Absorption Results ...................................................... 58
Metabolism Results ...................................................... 59
Literature Cited ................................................................... 63
Chapter 4. Summary and Conclusions ............................................... 82

vi

LIST OF TABLES

Chapter 2. Metolachlor Efficacy as Influenced by Three Acetolactate Synthase
Inhibiting Herbicides Under Field and Greenhouse Conditions

Table 1- Metolachlor, flumetsulam and halosulfuron plus furilazole treatments in

greenhouse studies ............................................................. 33
Table 2- Field site descriptions for corn and soybeans ........................... 34
Table 3- Bamyardgrass measurements 21 DAPL .................................. 35

Table 4- lncidences of antagonism of barnyardgrass control from
metolachlor: flumetsulam and metolachlorzhalosulfuron

combinations .................................................................. 36
Table 5- 1994 giant foxtail control in corn .......................................... 37
Table 6-1994 giant foxtail control in soybean ....................................... 38
Table 7-1995 giant foxtail control in corn ........................................... 39
Table 8-1995 giant foxtail control in soybean ....................................... 40
Table 9-1996 giant foxtail control in corn ........................................... 41
Table 10-1996 giant foxtail control in soybean ..................................... 42

Chapter 3. Physiological Basis for the Antagonistic Response of Giant Foxtail
(Setan'a faben' L.) to Metolachlor in the Presence of Flumetsulam,
Halosulfuron and Chlorimuron-ethyl.

Table 1- Reduction in giant foxtail shoot height dry weight by metolachlor,
chlorimuron, flumetsulam and halosulfuron with and without furilazole 28
days after treatment ............................................................. 65

Table 2- Giant foxtail shoot length as influenced by metolachlor combinations with

flumetsulam or chlorimuron when flumetsulam and chlorimuron were
applied 24 h prior to metolachlor ............................................ 66

vii

Table 3- Giant foxtail shoot length as influenced by metolachlor combinations with
halosulfuron with and without furilazole in the presence and absence of
fertilizer when halosulfuron was applied 24 h prior to metolachlor. . .67

Table 4- Site of uptake in giant foxtail for metolachlor, flumetsulam, chlorimuron
and halosulfuron with and without furilazole ............................. 68

Table 5- Relative soil mobilities of metolachlor, flumetsulam and chlorimuron
on soil thin-layer chromatography plates ................................. 69

Table 6- Relative soil mobilities of metolachlor, flumetsulam, chlorimuron and
halosulfuron with and without the safener furilazole as compared on soil
columns ......................................................................... 70

Table 7- Giant foxtail absorption of metolachlor 8 h after pretreatrnents of

metolachlor, an ALS inhibitor or a combination of metolachlor plus an
ALS inhibitor ................................................................. 71

viii

LIST OF FIGURES

Chapter 3. Physiological Basis of Giant Foxtail (Setan'a faberi L.) Response to
Metolachlor in the presence of Flumetsulam, Halosulfuron and
Chlorimuron-ethyl.

Figure 1- Radicle and shoot emergence of giant foxtail ........................... 73

Figure 2- Absorption of metolachlor by giant foxtail as influenced by ALS-
inhibiting herbicides plus metolachlor pretreatments .................. 75

Figure 3- Absorption of metolachlor by giant foxtail as influenced by ALS-
inhibiting herbicide pretreatments ........................................ 77

Figure 4- Metabolism of metolachlor by giant foxtail as influenced by ALS-
inhibiting herbicides plus metolachlor pretreatments .................. 79

Figure 5- Metabolism of metolachlor by giant foxtail as influenced by ALS-
inhibiting herbicide pretreatments ........................................ 81

ix

INTRODUCTION

Metolachlor is an acetanilide herbicide that is applied for grass control in both corn
and soybeans. Flumetsulam and halosulfuron are ALS-inhibiting herbicides that control
predominantly broad-[caved weeds and can be applied in combination with metolachlor for
broad spectrum weed control. In corn field trials in 1992 and 1993, a decrease in grass
control was observed where metolachlor plus flumetsulam or halosulfuron was applied
compared to metolachlor alone. A reduction in weed control could have a negative
environmental and economic impact and would be unacceptable to producers.

Antagonism of weed control from combinations of postemergence herbicides is a
well characterized interaction. Conversely, herbicides applied directly to the soil should
rapidly disassociate and act in an independent manner. It has always been assumed that the
risk for antagonism between preemergence herbicides was minimal. The appearance of thi 3
interaction has provided the opportunity to study preemergence antagonism more
thoroughly.

The goals of this research were to confirm the presence of this interaction under
more controlled conditions and to determine if this interaction with metolachlor was
isolated to flumetsulam and halosulfuron. If antagonism could be observed under more
controlled conditions, then experiments to characterized the nature of the interaction and
the basis for antagonism would be initiated. A decrease in grass activity from metolachlor
in the presence of an ALS-inhibiting herbicide has implications from the aspect of

potentially safening graminaceous crops against acetanilide herbicides.

Chapter 1

LITERATURE REVIEW

Metolachlor (2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)
aetamide) is an acetanilide herbicide that is applied for the control of grasses, yellow
nutsedge (cyperus esculentus L.) , and some broadleaf weeds in both corn (Zea mays L.)
and soybeans (Glycine max). Herbicides in this family are considered to be general
growth inhibitors affecting both root and shoot growth after seedling emergence. The
mode of action of this group of herbicides has never been satisfactorily elucidated though
it is suspected that these chemicals interfere with lipid synthesis through the disruption of
protein synthesis (Narsaiah and Harvey 1977, Chang and Merkle 1982).

ALS inhibitors include the imidazolinone, sulfonylurea, and triazolopyrimidine
sulfonailide classes of herbicides. DowElanco Inc. introduced a preemergence ALS
inhibiting herbicide in 1995, flumetsulam (N-(2,6-difluorophentyl)-5-methyl[1,2,4]triazolo
[1,5-a]pyrimidine-2-sulfonamide), that is sold only in premixture combinations. One of
these combinations is flumetsulam plus metolachlor. Preliminary field trials in corn at
Michigan State University in 1992 and 1993 documented a reduction in annual grass
control when flumetsulam or halosulfuron (methyl 3-chloro—5-(4,6-dimethoxypyrimidin-2-
ylcarbamoylsulfamoyl)-l-methylpyrazole-4-carboxylate) with and without the safener
furilazole (3-dichloroacetyl-5-(2-furanyl)-2,2-dimethyl-oxazolidine) was applied with
metolachlor when compared to metolachlor alone. The antagonsim in grass control could

not be linked to soil or environmental factors and could not be predicted with any

certainty. A reduction in grass control would be unacceptable to growers. Therefore,
characterization of this phenomena would be beneficial. This paper will provide a review
of the literature on metolachlor, flumetsulam, chlorimuron—ethyl (2-[[[[94-chloro—6-
methoxy-Z-pyrimidinyl) amino]carbonyl]amino]sulfonyl]benzoic acid), halosulfuron and

furilazole and their potential interactions.

METOLACHLOR

Metolachlor was released worldwide in 1974 by Ciba-Geigy under the trade name
of Dual“. The mode of action of metolachlor has never been satisfactorily elucidated.
Studies have indicated that the mode of action of the chloroacetanilide herbicides is the
inhibition of protein synthesis or related systems (Deal and Hess 1980). Metolachlor,
propachlor (2-chloro-N-(l-methylethyl)-N-phenylacetamide), and alachlor ( 2-chloro-N-(2,
6-diethylphenyl)-N-(methoxymethyl)acetamide) are known to inhibit cell division in oat
(A vena fatua L.) root and CDAA (2-chloro-N,N-di-2-propenylacetamide) and propachlor
are known to inhibit protein synthesis in vivo (Narsaiah and Harvey 1977, Deal et a1.
1980, Chang and Merkle 1982). Propachlor and alachlor have also been shown to inhibit
gibberellic acid induced production of protease and alpha-amylase (Chang and Merkle
1982). Though implicated in the synthesis of proteins, metolachlor did not effect the
assembly of ribosomal subunits onto mRN A and other linked processes (Chang and Merkle
1982) thereby confounding previous findings. Another proposed mechanism for the action
of these herbicides is the alteration of plant membranes and lipid synthesis.

The effects of metolachlor on lipid synthesis are unclear. Leucine incorporation

was reduced in corn and barley when exposed to high concentrations of metolachlor (Pillai
et al. 1979). In cucumbers, metolachlor and alachlor decreased l“C-leucine incorporation
during protein synthesis (Deal and Hess 1980). Fungal and bacterial cultures were
observed in nutrient solutions containing 1.0 x 104 M of metolachlor indicating the
possibility of leakage of a bacterial of fungal growth component through a perforated
membrane (Mellis et al. 1982). Leakage of P32 from onion root tips suggests that the loss
of membrane integrity is a factor in the mode of action of these herbicides. This membrane
leakage was also observed with the acetanilide CDAA where lipogenesis was inhibited due
to decreased malonic acid incorporation into membrane lipids (Mann et al. 1965). Lipid
synthesis in cotton root tips was 26% lower in the presence of both metolachlor and
alachlor. Specifically, there was a decrease in phospholipid and phosphotydlcholine
synthesis (Mellis et al. 1982). This decrease in choline incorporation into the phospholipid
phosphatidyl-choline in the presence of acetanilides has been observed in other
experiments (Deal et al. 1980).These data support interference with lipid synthesis as a
possible mode of action for metolachlor.

Sorption of metolachlor to soil particles is best described by a Freundlich isotherm
(Obrigawitch et al. 1981). Adsorption was correlated to soil clay content and organic
carbon content (Braverman et al. 1986) and multiple regression analysis suggested both
soil factors play a role in alachlor and metolachlor sorption (Peter and Weber 1985).
Freundlich adsorption constants for five soils were highly correlated to percent clay and
organic carbon (R2 = 0.99 and 0.94, respectively) (Braverman et al. 1986). Absorption

of metolachlor was pH dependent. Sorption was influenced by location in the soil profile

(Bouchard et al. 1982). Soil from the 10 to 20 cm range adsorbed more herbicide
compared to the soil obtained 40 to 50 cm depth. This could be a result of lower organic
matter content of the soil at this depth. Previous research indicated that metolachlor sorbs
to organic matter (Weber and Peter 1982, Pusino et al. 1992). It has been suggested that
the carbonyl oxygen of metolachlor forms a dipole-ion bond with adsorbed cations and H -
bonds with water molecules and carboxyl and hydroxyl groups on organic matter.
Metolachlor is present as a neutral molecule in most soil systems though uneven
distribution of electrons can cause slight polarization (Bouchard et al. 1982).Therefore,
metolachlor should be considered a moderately persistent compound. Acetanilides have
been found in soils at low levels for up to two years after application (Bouchard et al.
1982, Braverman et al. 1986). Herbicide concentration did not effect the degradation rate
though temperature positively correlated to increased degradation. CO; levels were not
affected by metolachlor concentration indicating that metolachlor did not inhibit microbial
activity. The degradation of metolachlor was not significantly correlated with percent
moisture or CO, concentration but was correlated to time. Sterilized soil showed no
significant breakdown of metolachlor after four months. Therefore, microbial breakdown
is the predominant mode of metolachlor degradation in a soil system.
Microbial breakdown of metolachlor occurs predominantly through dechlorination
(Liu et al. 1989, McGahen et al. 1978), hydroxylation (Liu et al. 1991), or demeth ylation
(Saxena et al. 1987). A mixed bacterial culture was shown to accumulate, and
metabolically, transform metolachlor more than any one microbe alone. The mechanism

of binding and accumulation was not elucidated (Liu et al. 1989). This could be the result

of a synergistic interaction of the various microflora which leads to enhanced microbe
growth and vigor. Successive cultures of the soil microbes belonging to the species
Fusarium were able to metabolize metolachlor with increasingly alacrity suggesting an
enzymatically induced system of transformation (Saxena et al. 1987).

Leaching was an important factor in metolachlor dissipation. Half life increased
when metolachlor dissipation was dependent exclusively on degradation (Braverman et al.
1986). Movement of metolachlor decreased over time possibly due to the differences in
bonding energies between the metolachlor sorbed to interlayer surfaces and that sorbed to
multi-molecular layers of clay. Pusino postulated that organic matter in close association
with the clay fraction is excluded from binding metolachlor (Pusino et al. 1989). Clay and
humic Kd values were lower than would be expected if the two soil components acted
independently with respect to metolachlor binding. Clay/organic matter associations with
metolachlor decreased sorption but increase the soil mobility of metolachlor.

Metolachlor undergoes rapid glutathione conjugation followed by breakdown to
a cysteine conjugate in maize and rice (Cole and Owen 1987). The potential for rice
(Oryza sativa L.) injury with metolachlor is increased when the field is flooded
(Braverman et al. 1986). Hatzios (1983) found that the detoxification of metolachlor in
plants was dependent on molecular oxygen and potentially dependent on a mixed function
oxidase system. ABT (l-aminobenzotriazole) acts as a mechanism-based inactivator for the
mixed function oxidase dependent on cytochrome P-450 (Cole and Owen 1987). It inhibits
cinnamic acid-4-hydroxylase in plants and mammals. 2,4-DP (3(2,4-dichlorophenoxy)-l-

propyne) inhibits hepatic but not plant P-450. These compounds could modify the action

of herbicides that are oxidatively metabolized. ABT did not effect metolachlor metabolism
in corn but decreased metabolism in cotton (Gossypium hirsutum) (Cole and Owen 1987).
Metabolism was stimulated in corn upon the addition of 2,4-DP. Evidence for cytochrome
P450 involvement in metolachlor metabolism is Still conjectural.

The site of uptake for metolachlor was the shoot in corn and barley (Hordeum
vulgare L.) as evidenced by a reduction in corn and barley heights.(Pillai et al. 1979).
Applications of metolachlor to root or seed zones did not effect corn or barley height but
did appear to be the method of entry in pea (Pisium sativum L. ’Thomas Laxton’) (Pillai
et al. 1979). The reason given for this difference in sensitivities between the monocots
and the dicots was that the primary root meristem of dicots is below the seed level while
the primary roots and adventious root meristems for monocots are above the seed.

Absorption and metabolism have been implicated as the basis for differences in
tolerance between plant species to acetanilide herbicides (Hamill and Penner 1973).
Selectivity has also been shown to be related to the rate of metolachlor metabolism.
Differences in tolerance to metolachlor have been observed between corn varieties that
exhibit differential metabolism with the susceptible varieties detoxifying metolachlor more
slowly (Cottingham and Hatzois 1992). The susceptible species yellow nutsedge absorbed
metolachlor faster and to a greater extent than the tolerant species corn (Cottingham and
Hatzios 1992). Hamill and Penner (1973) found that more alachlor was absorbed by
barley, which is a susceptible species, compared to corn which is tolerant. Corn
metabolized the metolachlor more rapidly than barley and rapid metabolism appeared to

be the primary reason for greater tolerance in corn.

Increased metolachlor metabolism was linked to higher levels of reduced
glutathione and/or to expression of a glutathione S-transferase isozyme that possessed
greater herbicide specificity (Cottingham and Hatzios 1992). This was not true for other
herbicides detoxified through the same pathway. When atrazine(6-chloro-N-ethyl-N’-(l-
methylethyl)-1,3,5-triazine-2,4-diamine) was applied to velvetleaf (Abutilon theophrasti
Medicus), the amount of glutathione was not significantly different between resistant and
susceptible biotypes (Anderson and Gronwald 1991). The most glutathione and most
glutathione-S-transferase activity was in leaf tissue regardless of biotype. Glutathione-S-
transferase activity was 40 and 25 % greater in the susceptible biotype leaves and stems,
respectively, compared to the resistant variety when l-chloro—2,4-dinitrobenzene was used
as a substrate. When atrazine was used as a substrate, the level of glutathione-S-transferase

activity was approximately four times greater in leaf and stem tissue of the resistant

biotype.

FLUMETSULAM
The triazolopyrimide sulfonanlide, flumetsulam, is an ALS inhibiting herbicide
produced by DowElanco for preemergence broadleaf weed control in corn and soybean.
Flumetsulam has a water solubility that increases with increasing soil pH and is considered
to be a weak acid (pK, of 4.6) (Kleschick et al. 1992). The estimated half life for
flumetsulam in aerobic soils ranged from 2 weeks to 4 months and was 183 days in a
highly anaerobic system (W olt et al. 1992). ALS inhibitors, such as flumetsula m, disrupt

the production of the branch chain amino acids valine, leucine and isoleucine (Upchurch

et al. 1962, Fontaine et al. 1991, Kleschick et al. 1992).

Leucine is a competitive inhibitor with ALS- inhibiting herbicides for the binding
Site on the ALS enzyme (Subramanian et al. 1991). These herbicides are suspected to bind
to the regulatory site of the enzyme. Both leucine and valine, but not isoleucine, are
feedback inhibitors of ALS and resistant varieties of plants were resistant to feedback
inhibition to varying degrees (Subrarnanian et al. 1991). Almost all cases of naturally
occurring resistance to an ALS inhibitor has been due to an altered ALS enzyme
(Schmitzer et al. 1993). Cross resistance varied among the three major classes of ALS
inhibitors suggesting overlapping but not identical binding sites. Resistance was correlated
with the ability of a plant to rapidly metabolize the herbicide and reduce the concen tration
in the plant below that required for growth inhibition (J aworski 1969).

The selectivity of flumetsulam is due to differences in rates of oxidative metabolis m
and detoxification (Frear et al. 1993). Metabolism was more rapid in wheat (Triticum
aestivum L.) leaves compared to corn and barley, even though the metabolic pathways fo r
all three plants were similar. Three metabolites were found in these plants as well as the
glucoside conjugates of those metabolites. The metabolites were designated Ml , M2, and
M3 based on the increase in polarity in thin-layer chromatography experiments. M1 is
hydroxylated on the 4-C of the 2,6—difluro-ring and M2 is hydroxlated on the methyl group
of the pyrimidine ring. The third metabolite is hydroxylated in both positions and both M1
and M2 can convert to the M3, form. Differences in estimated Km values for M1 and M2
formation suggests the presence of several forms of inducible cytochrome P450 with

varying degrees of sensitivity to the substrate flumetsulam (Frear et al. 1993). This

experiment showed that flumetsulam hydroxylase required NADPH and molecular oxygen
to function and metabolite production was inhibited by cytochrome P450 inhibitors.
Therefore, flumetsulam metabolism is dependent on mixed function oxidase activity.

In strongly anaerobic systems, flumetsulam degraded almost totally to reduced
flumetsulam hydrate (Wolt et al. 1992). This product was reverted back to flumetsulam
upon aeration. Under anaerobic and aerobic conditions, metabolism and degradation were
of a first order kinetic reaction. There is a minimal possibility that this metabolite would
occur in production agriculture systems. The only place where a build-up of reduced

flumetsulam hydrate would be of concern is in a static, strongly anaerobic environment.

CHLORIMURON-ETHYL

Chlorimuron-ethyl is a sulfonylurea herbicide produced by Dupont and sold under
the trade name of Classic. Chlorimuron is used for broadleaf weed control in soybean.
This herbicide is applied postemergence or in prepackaged mixtures with metribuzin and
sulfentrazone and sold as Canopy and Authority / Canopy XL, respectively, for
preemergence weed control. Like flumetsulam, chlorimuron belongs to the ALS inhibitor
class of herbicides (Herbicide Handbook 1994). Preemergence weed control with
chlorimuron is becoming more common since the recent marketing of these prepackaged
mixtures which include chlorimuron.

The efficacy of a soil applied herbicides is dependent upon soil factors and
herbicide bioavailability (Goetz et al. 1989). The mitigating factor in the availability of

most herbicides in a soil system is the soil organic matter content (Weber 1970, Vencill

10

and Banks 1994). As with other sulfonylurea herbicides, chlorimuron is a weak acid with
a pKa of 4.2 and a water solubility that increases with increasing pH (Herbicide Handbook
1994). At soil pH values between 5.5 to 8, the predominant State of chlorimuron would
be anionic. This results in sorption being highly dependent on the total concentration soil
constituents such as clay, ferrous oxides and positively charged functional groups on the
organic matter moiety. Goetz et a1. (1989) found that chlorimuron sorption to soil was
dependent on pH with sorption increasing as soil pH decreased. They attributed the
increase in chlorimuron sorption to the iron and aluminum oxide concentration in the soil.
The zero net point charge for these molecules would have been positive in the pH range
of the experiment and decreased as the pH increased. They concluded that iron and
aluminum oxides, clays and organic matter were available for chlorimuron sorption.

Bioavailability of chlorimuron was time dependent (V encill and Banks 1994).
Chlorimuron dissipation from the soil was initially rapid and decreased over time.
Sicklepod (Sida spinosa L.) root length was used as an indicator of the presence of
chlorimuron. After 120 days, less than 5% of the original field use rate was detectable.
The time for 50% dissipation of bioavailable chlorimuron CDT”) was positively correlated
to precipitation and temperature.

Plant absorption of soil applied chlorimuron is not well documented. Yellow and
purple nutsedge (cyperus rotundus L.) absorbed chlorimuron through roots, Shoots and
tubers (Reddy and Bendixen 1989). Chlorimuron absorbed by nutsedge tubers was not
translocated while chlorimuron absorbed through any other structure resulted in

translocation into shoot tissue. Symplastic movement, as influenced by transpiration rate,

11

appeared to be the cause of any herbicide movement out of the treated leaf. Transpiration
was also implicated as the driving force of absorption and translocation in these two
nutsedge species. A reduction in transpiration following an application of the ALS-
inhibiting herbicides primisulfuron (2-[[[[[4,6-bis(difluoromethoxy)-2-
pyrimidinyl]amino]carbonyl]amino]sulfonyllbenzoic acid) and nicosulfuron (2-[[[[(4,6-
dimethoxy-Z-pyrimidinyl)amino]carbonyl]amino] sulfonyl]-N, N-dimethyl-3-pyridine
carboxamide) has been documented in sugarbeet (Beta vulgaris L.) (Novosel 1995). These
data imply that absorption and translocation of chlorimuron could be reduced if
transpiration was reduced.

Chlorimuron reduced the efficacy of the ACCase-inhibiting graminicide quizalofop
(( + /-)-2-[4-[(6-chloro-2-quinoxalinyl)oxy]phenoxy]propionic acid) (Bjelk and Monaco
1992). Fatty acid biosynthesis, the site of action of quizalofop, has been linked to the
concentration of valine, leucine and isoleucine in the plant (Murphy and Sturhpf 1981).
Since the synthesis pathways of these three amino acids were inhibited by the ALS
inhibitors, studies were initiated to investigate the effect of chlorimuron on fatty acid
biosynthesis using l“C-pyruvate and l“C-acetate incorporation as an indicator (Bjelk and
Monaco 1992). Chlorimuron had no effect on l“C-acetate incorporation compared to
quizalofop (Bjelk and Monaco 1992). Excess pyruvate did not alleviate the effects of
quizalofop indicating that the basis of the antagonism is not the increased availability of
the common substrate pyruvate. It was therefore concluded that the basis for the observed

antagonism was not due to the disruption of fatty acidbiosynthesis by chlorimuron.

12

HALOSULFURON ANS FURILAZOLE

Halosulfuron, previously MON 12000, is a new ALS inhibiting herbicide produced
by Monsanto. Furilazole, previously MON 13900, is a safener added to halosulfuron.
Halosulfuron without the safener is sold as Pemtit“. Originally, the combination of
halosulfuron plus furilazole was to be sold under the trade name of Battalion“ but this
product has been discontinued. Halosulfuron is sold for postemergence yellow nutsedge
and broad-leaved weed control in corn and is labeled for preemergence use on
imidazolinone resistant corn. Halosulfuron and furilazole have low mammalian toxicity
(Anonymous 1993). There is very little information published with respect to halosulfuron.
Most of the published literature focused on furilazole.

Uptake of furilazole occurs predominantly through shoot interception in broad-
leaved weeds though some root activity has been noted (Bussler et al. 1991). Conversely,
root interception is the primary route of entry for halosulfuron in broad-leaved plants
(Herbicide Handbook 1994). The safening effect of furilazole resulted from an increased
rate of metabolism of halosulfuron though the mechanism is not clearly understood
(Anonymous 1993). It was suspected that the mechanism involves mixed function oxidase
activity.

The metabolic pathway for furilazole in corn and sorghum is conversion to oxamic
acid and/or an alcohol followed by conjugation to an alcohol glucoside (Bussler et al.
1991, Anonymous 1993). Oximes have the potential to reactivate some hydrolytic enzyme S
(Chang and Merkle 1982). They can act as a nucleophilic reagent that reacts with the

phosphorous on the enzyme inhibitors thereby disrupting the phosphorous-enzyme bond.

13

The primary metabolite of furilazole appears to be glucose/fructose conjugate with no
parent material present in plant material after 24 h. Furilazole increased the metabolism
of NC-3l9, a sulfonylurea herbicide, in corn but did not affect the expression of ALS
activity (Bussler et al. 1991). Furilazole also is a safener to acetanilide herbicides. This
is most likely due to an increase in glutathione conjugation as a result of glutathione
induction.
INTERACTIONS

The use of herbicide combinations is a common weed control practice in
many crop and non-crop situations. Herbicide mixtures may result in several different
interactions that are categorized as additive, synergistic, and antagonistic (Hamill and
Penner 1973, Akobundu et al. 1975) .

A synergistic interaction could increase crop production capabilities by reducing
weed competition (Winkle et al. 1981). In greenhouse studies, soil applied alachlor acted
synergistically with imazaquin (2-[4,5-dihydro—4-methyl-4-(l-methylethyl)-5-oxo- 1H-
imadazol-2-yl]-3quinolinecarboxilic acid) and DPX-F6025 (chlorimuron) when applied to
1 to 3 leaf stage Sicklepod (Edmund et al. 1987). The mechanism of this interaction was
not elucidated. The synergism observed between atrazine and alachlor was not due to any
direct effect on photosynthesis (Akobundu et al. 1975) but rather due to a reduction in
chloroplast protein and protein synthesis. Metabolism of atrazine was not effected by the
addition of alachlor which indicates that alterations in metabolic functions were not
responsible for the observed synergism.

Antagonism of grass control with tank mixtures of acetanilide herbicides and ALS

14

inhibiting herbicides was observed in field trials (Kells 1992, Owen 1993, Simmons 1993,
Lueshen 1995, Wax 1995). Incidences of decreased injury to grass species when
metolachlor or alachlor was applied with another herbicide is not limited to mixtures with
ALS inhibitors. Metolachlor and alachlor reduced the injury to corn from buthidazole (3-
[5-(1, l-dimethylethyl)-l , 3 ,4-thiadiazol-2-yl]-4hydroxy-1-methyl-2-imidazolinone) when
applied both preplant incorporated and preemergence (York and Slife 1981). While a
decrease in crop injury is rarely referred to as antagonism, this shows the effect of

acetanilide herbicides on grass Species in the presence of a tank mix partner.

HERBICIDE INTERACTIONS

Antagonized grass control with tank mixtures containing ALS inhibitors are not
limited to acetanilides. Graminicides that inhibit the enzyme acetyl-CoA carboxylase
(ACCase), like the aryloxyphenoxypropionates and sethoxydim (2-[1-(ethoxyimino)butyl]-
5-[2-(ethylthio)propyl]-3-hydroxy-2-cyclohexen-1-one), were antagonized when an
imidazolinone such as imazethapyr (2-[4,5-dihydro-4-methyl-4-(1-methlethyl)-5-oxo-1H-
imidazol-2-yl]-5-ethyl-3-pyridinecarboxylic acid) was applied (Castro-EScobar 1997).
Chlorimuron has been shown to reduce the efficacy of quizalofop on broadleaf signalgrass
(Bracharia platyphylla L.) (Bjelk and Monaco 1992). Chlorsulfuron (2-chloro-N-[[(4-
methoxy-6-methyl-l , 3 ,5-triazin-2-yl)amino]carbonyl]benzenesulfonamide) reduced the wild
oat control from diclofop (( +/-)-2-[4-(2,4-dichlorophenoxy)phenoxy]propionic acid) by
35 % in greenhouse studies performed by O'Sullivan (O’Sullivan and Kirkland 1984).

Sasaki, Konishi and Nagano (Sasaki et al. 1995) found that ACCase in plants evolved into

15

two different forms of the enzyme. The site of fatty acid synthesis in plants is the plastid
which is thought to be evolved from a prokaryotic symbiosis. Fatty acid synthase catalyzes
the reaction of 1 acetyl CoA + 7 malonyl CoA to create 16-C fatty acids. Prokaryotic
organisms and plants have a type of fatty acid synthase called type 11 while eukaryotic
organisms have type I. ACCase supplies malonyl CoA to the fatty acid synthesis pathway.
ACCase also has prokaryotic and eukaryotic forms. Dicots contain both types of enzymes
(eukaryotic in the cytosol and prokaryotic in the plastids) and grasses contain only type I.
This could explain why grasses are susceptible to sethoxydirn and
aryloxyphenoxypropionates. These herbicides target plastid ACCase, therefore prokaryotic
ACCase would not be susceptible while the eukaryotic form would be susceptible. The
discovery of different forms of ACCase leads to questions as to the effects of the ALS
inhibitors on these different isozymes and how grass response to tank mixture

combinations is influenced.

16

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Anonymous. 1993. Mon 12000 technical data sheet. Monsanto publication, Monsanto
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19

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20

21

Chapter 2
METOLACHLOR EFFICACY AS INFLUENCED BY THREE
ACETOLACTATE SYNTHASE INHIBITING HERBICIDES UNDER
FIELD AND GREENHOUSE CONDITIONS
Abstract. In preliminary field trials, a reduction in giant foxtail control was observed
when flumetsulam or halosulfuron was applied with metolachlor compared to metolachlor
alone. Greenhouse studies were initiated to study potential interactions between
acetolactate synthase (ALS) inhibiting herbicides and metolachlor. Metolachlor at 0.14,
0.28, and 0.56 kg ai ha" was applied alone and in combination with flumetsulam at 0.007 3
and 0.015 kg ai ha" and halosulfuron at 0.011 and 0.021 kg ai ha". Flumetsulam and
halosulfuron alone provided 13 to 23% barnyardgrass control and metolachlor at 0.14 kg
ha'1 provided 82 % control. Combining flumetsulam and halosulfuron with metolachlor did
not increase herbicide activity beyond that observed from metolachlor alone regardless of
the parameter evaluated. Combinations of 0.14 kg ha" of metolachlor plus 0.011 and
0.021 kg ha‘1 of halosulfuron or 0.015 kg ha'1 of flumetsulam resulted in antagonism of
barnyardgrass control as measured by visual control, grass height and plant dry weight
according to Colby’s multiplicative interactive model. Visual control and plant dry weight
were also antagonized when 0.028 kg ha'1 of metolachlor was applied with 0.021 kg ha‘1
of halosulfuron. Field studies were conducted for 3 years in corn and soybean to evaluate
giant foxtail control from metolachlor alone and in tank mixtures with flumetsulam,
halosulfuron, and chlorimuron. In 1994 a year with limited rainfall, the addition of an
ALS inhibiting herbicide to metolachlor neither increased nor decreased grass control in

soybeans compared to metolachlor alone. In corn, giant foxtail dry weight was decreased

more when flumetsulam or halosulfuron plus safener was applied with metolachlor at 0.5 6
kg ha ‘1 compared to metolachlor alone. In 1995 and 1996, years with adequate moisture,
giant foxtail control by metolachlor was not consistently enhanced or reduced when
applied with flumetsulam, halosulfuron, or chlorimuron. Thus, results from 3 years of
field trials showed neither an increase nor a decrease in grass control when these ALS-
inhibiting herbicides were applied in combination with metolachlor compared to

metolachlor alone.

INTRODUCTION

Metolachlor (2-chloro-N-(2-ethyl—6-methylphenyl)-N-(2-methoxy-1-methylethyl)
aetamide) is an acetanilide herbicide that is used for the control of annual grasses, yellow
nutsedge (cyperus esculentus L.), and some broad-leaved weeds in both corn (Zea mays)
and soybeans (Glycine max)(Herbicide Handbook 1994.). The acetolactate synthase (ALS)
inhibitors are a broad class of herbicides used in many cropping systems for selective weed
control (Herbicide Handbook 1994.). Flumetsulam (N-(2,6-difluorophenty1)-5-methyl
[1,2,4]triazolo [1,5-a]pyrimidine-2-su1fonamide), halosulfuron (methyl 3-chloro-5-(4,6-
dimethoxypyrimidin-Z-ylcarbamoylsulfamoyl)-l-methylpyrazole-4-carboxylate), and
chlorimuron (2-[[[[94-chloro-6-methoxy-2-pyrimidinyl)amino]carbonyl]amino]sulfonyl]

benzoic acid) are ALS- inhibiting herbicides that control many broad-leaved weeds.
Flumetsulam, halosulfuron with and without the safener furilazole [3-dichloro

acetyl-5-(2-furanyl)-2,2-dimethyl-oxazolidine], and chlorimuron alone have provided 30

22

to 70% giant foxtail control (Renner and Kells 1996). In years where the performance of
metolachlor alone was unacceptable, tank mixtures of metolachlor that include these ALS-
inhibiting herbicides could potentially increase grass control in an additive fashion.
However, in field trials at Michigan State University in 1992, annual grass control was
reduced when flumetsulam or halosulfuron with and without the safener furilazole was
applied with metolachlor when compared to metolachlor alone (Kells and Renner 1992).
A lack of an additive response could be a result of a decrease in the grass activity of eithe r
metolachlor or the ALS inhibitor. Characterization of both the additive and antagonistic
interactions of these herbicides would decrease the potential for additional herbicide input
for annual grass control in corn and soybeans as well as increase the understanding of
activity of these herbicides in grassy weed species. Therefore research was initiated to
determine the presence or absence of an additive response between metolachlor and the
three ALS-inhibiting herbicides flumetsulam, halosulfuron, and chlorimuron in both

greenhouse and field studies.

MATERIALS AND METHODS
Greenhouse study. Barnyardgrass (Echinochloa crus-galli L.) had 80 to 95 % emergence
and was used as the bioassay species. Twenty seeds were planted in 454 g pots contai ning
a loam soil with 3.3 % organic matter. The pots were prepared for subirrigation by drilling
holes equidistantly spaced on the bottom near the outer edge. Twenty treannents were

applied to the pots using an 8003B nozzle at 209 L ha'1 and 207 kPa (Table 1). Pots were

23

surface watered immediately after herbicide application and then blocked by replication.
All pots were uniformly watered; alternating between surface watering and subirrigation.
Pots within replicates were re-randomized on a weekly basis. Plant height (cm), number
of live plants emerged, and visual estimates of growth reduction were measured 7, 14, and
21 days after treatment (DAT) . Median height was recorded to the nearest 0.5 cm.
Emergence of a plant was counted when any green part was visible above the soil. Visual
evaluations compared the amount of biomass in a pot to that of the untreated control. All
above ground material was harvested 21 DAT, dried, and weighed to the nearest 0.1 mg
on a per pot basis.

The experiment was a randomized complete block with three replications and the
experiment was conducted twice. Since there was a main effect interaction, statistical
analysis was completed on the means of each experimental unit and subjected to F isher's
Protected LSD at the 0.05 significance level. Analysis of treatments showed no significant
interactions and therefore results are presented as the average of both experimental units.
Colby’s multiplicative interactive model was applied to the data to determine the presence
or absence of antagonism.

Field studies. Field studies were conducted at two sites in 1994, 1995, and 1996 at the
Agronomy farm at East Lansing, Michigan. Imidazolinone resistant corn was planted at
the first field site and Conrad soybeans planted at the second site. The field site
descriptions are described in Table 2. Corn was planted on May 10, 1994, May 8, 1995,
and May 17, 1996 and soybeans on May 15, 1994, May 16, 1995, and May 30, 1996. The

herbicide treatments were applied preemergence immediately following planting. A

24

postemergence application of bentazon (3-(l-methylethyl)-(l H)-2,1,3-benzothiadiazin-4(3
H)-one 2,2-dioxide) was made in 1994 and 1995 in the metolachlor alone treatments for
broadleaf weed control. In 1996, treatments were hand weeded to control broad-leaved
weeds. The corn study consisted of sixteen treatments including metolachlor at 2.24, 1.12 ,
and 0.56 kg ai ha", halosulfuron with and without safener at 0.084 kg ai ha‘l and
flumetsulam at 0.063 kg ai ha". Each herbicide was applied alone and metolachlor was
also applied in a tank mixtures with flumetsulam and with halosulfuron. In 1994, the
soybean study consisted of twenty-one treannents including metolachlor at 2.24, 1.68,
1.12, 0.84, and 0.56 kg ha"; flumetsulam at 0.063 kg ha'1 and chlorimuron at 0.031 kg
ai ha". These herbicides were applied alone and in a tank mixture combinations of
metolachlor plus an ALS inhibitor. In addition, the prepackaged mixture of flumetsulam
plus metolachlor was applied at five rates which corresponded to the metolachlor rates in
the tank mix combinations. The rate of flumetsulam decreased in the premixture as the
metolachlor rate decreased while the flumetsulam rate remained constant in the non-premix
tank mixtures. In soybeans in 1995 and 1996, the 2.24 kg ha" rate of metolachlor was
changed to 2.1 kg ha'1 and 2.35 kg ha", respectively due to differences in soil type and
percent organic matter at the field sites. The average density of giant foxtail (Setaria faberi
L.) per 0.5 m2 was 52 and 70 plants per quadrat in corn and soybean, respectively. Giant
foxtail control was evaluated visually 28 and 56 days after planting (DA PL). Two 0.5 m2
quadrats were harvested from each plot 56 DAPL and giant foxtail number per quadrat,
total plant dry weight per quadrat, and average weight per plant were determined.

The experimental design was a randomized complete block with three replications

25

in corn each year. There were three replications in soybean in 1994 and four replications
in 1995 and 1996. Statistical analysis of field data showed a year by treatment interactio n,
therefore results are presented separately for each year. Means were separated by Fisher' 5

protected LSD at the 0.05 level of Significance.

RESULTS AND DISCUSSION

Greenhouse studies. Flumetsulam and halosulfuron alone provided 13 to 23 % visual
control of barnyardgrass and Significantly decreased plant dry weight (Table 3).
Barnyardgrass control was reduced compared to metolachlor alone when 0.015 kg ha‘1 of
flumetsulam was added to 0.14 kg ha‘1 of metolachlor as reflected in greater plant height,
emergence and dry weight (Table 3). This response was determined to be antagonistic
according to Colby’s multiplicative interactive model (Table 4). Antagonism of emergence
occurred following applications of flumetsulam at 0.0073 kg ha'1 plus metolachlor at 0.14
and 0.28 kg ha '1. Additionally, a decrease in visible grass control and an increase in giant
foxtail height and dry weight was observed when halosulfuron at 0.011 and 0.021 kg ha"
was applied with metolachlor at 0.14 kg ha‘l though no effect on emergence was apparent.
Halosulfuron at 0.021 kg ha‘1 in combination with 0.28 kg ha'1 of metolachlor antagonized
visual control of barnyardgrass as well as emergence and plant dry weight. Synergism was
not evident for any measurement with any treatment.

Field Studies. W Grass control with 2.24 kg ha" of metolachlor was 67 %

28 DAPL in corn while flumetsulam alone gave 10% grass control (Table 5). Only 0.56

26

cm of rain fell between May 10 and May 30, 1994 which was insufficient for herbicide
activation. From June 6 to July 7, over 25 cm of rain fell with an additional 10 cm of
rainfall in July and 14 cm in August. In 1994, a year with little rainfall early in the season ,
the ALS-inhibiting herbicides did not posses grass activity in spite of their greater
solubility compared to metolachlor.

Halosulfuron without safener and flumetsulam did not significantly increase gras 3
control in corn when combined with any rate of metolachlor 28 DAPL. Variability of
visual data decreased from 25% 28 DAT to 14% 56 DAT. By 56 DAPL, metolachlor at
2.24 kg ha" provided 65% visible control of annual grasses and reduced grass number
and dry weight by 50%. The addition of halosulfuron with safener or flumetsulam to
metolachlor at 0.56 kg ha'1 increased visible grass control compared to metolachlor alone
56 DAPL (Table 5). This was not reflected in a decrease in the number of grass per
quadrat but rather in dry weight per plant when compared to metolachlor al one (Table 5).

Metolachlor alone gave slightly greater grass control in soybean compared to corn
due to 3 to 4 cm of irrigation applied 17 DAPL (Table 6). Giant foxta i1 control 28 DAPL
in soybean was 80% with 1.12 kg ha'1 of metolachlor and only 33 % when 0.063 kg ha'1
of flumetsulam was added. By 56 DAPL, differences between these two treannents was
no longer evident. Chlorimuron alone provided 67 % grass control 28 DAPL as did
metolachlor at 0.56 kg ha'1 plus chlorimuron (Table 6). Poor grass control was observed
where flumetsulam was applied with 0.56 kg ha'1 of metolachlor in a tank mixture and
with the premix applications 56 DAPL.

Metolachlor, flumetsulam and chlorimuron alone reduced the total number of grass

27

per quadrat but not plant dry weight (Table 6). Combining metolachlor with an ALS
inhibitor did not reduce plant number compared to metolachlor or the ALS inhibitor alone.
Plant dry weight was not increased compared to metolachlor alone except with metolachlor
at 2.24 kg ha‘1 plus chlorimuron which was not reflected in grass number or visual ratings
(Table 6).

W In 1995, 5.1 cm of rain fell between May 8 and June 5 and
metolachlor alone at 0.56 kg ha'1 provided 75% grass connol in com 28 DAPL (Table 7).
Halosulfuron provided 40 to 47% grass control and flumetsulam provided 81% grass
control. Tank mixtures of flumetsulam or halosulfuron with metolachlor did not
significantly enhance or reduce visual grass control, grass number, or plant dry weight
compared to metolachlor alone (Table 7).

Giant foxtail control was 98% from 0.56 kg ha'1 of metolachlor 28 DAPL in
soybean (Table 8). Flumetsulam provided 62% grass control and chlorimuron provided
34 % control 28 DAPL. The activity of chlorimuron on giant foxtail should have been
greater than in 1994 since there was adequate moisture for herbicide activation in 1995 and
not in 1994. The reasons for the lower grass activity of chlorimuron in 1995 are still
unclear.

Chlorimuron added to 1.12 kg ha'l of metolachlor gave significantly lower grass
control compared to metolachlor alone 56 DAPL but this was not evident in increased
plant number or plant dry weight (Table 8). Metolachlor, flumetsulam and chlorimuron
alone reduced grass numbers and combinations did not increase or decrease control

compared to metolachlor alone. Plant dry weight was increased with the tank mixture of

28

flumetsulam plus metolachlor at 0.84, 1.12 and 2.1 kg ha" and with the prepackage
mixture containing 2.1 kg ha'1 of metolachlor. This response was not observed in other
measurements.

W Corn and soybean were planted later in 1996 due to cold, wet spring
conditions. Rainfall following herbicide application was over 5 cm in June and metolachlor
alone at 0.56 kg ha'1 provided 96% grass control in corn (Table 9). The ALS inhibitors
alone provided 67 to 78 % grass control in corn while tank mixtures with metolachlor gave
greater than 90 % control 56 DAPL. Metolachlor, flumetsulam and halosulfuron with and
without safener reduced the number of grass and reduced plant dry weight compared to
the untreated control (Table 9). Metolachlor alone at 0.56 kg ha‘1 reduced the number of
giant foxtail by 93% and foxtail dry weight by 82%. With the exception of 1.12 kg ha",
metolachlor applied with halosulfuron plus safener showed no significant effect on weight
per plant with tank mix combinations (Table 9).

Metolachlor alone at 0.56 kg ha" provided 90% giant foxtail control and
flumetsulam and chlorimuron provided 45 % and 65 % control, respectively, in soybeans
28 DAPL (Table 10) . By 56 DAPL, metolachlor alone at 0.56 kg ha“ provided 90% grass
control compared to 72 % from the prepackaged mixture containing the equivalent
metolachlor. Average plant dry weight in this premixture application was greater than an
equivalent rate of metolachlor alone though the number of foxtail per quadrat did not
significantly increase (Table 10).

Combinations of metolachlor at 0.14 kg ha'l and flumetsulam at 0.015 kg ha" or

halosulfuron at 0.011 and 0.021 kg ha'1 resulted in reduced grass control in greenhouse

29

experiments. These interactions were determined to be antagonistic according to Colby’s
multiplicative model. However, reduced grass control did not occur consistently in 3 years
of field trials in corn and soybeans. The ratio of metolachlor to ALS inhibitor may be
important in the prediction of antagonism in the greenhouse as more incidence of
antagonism occurred when the ratio of metolachlor to ALS inhibitor was lower.
Antagonism with the commercial premix ratio of metolachlor / flumetsulam (38:1) was
observed in a singular incidence in the field. However, ratios of metolachlor at 0.56 kg
ha’1 to flumetsulam, halosulfuron and chlorimuron were 9: 1, 7 :1 and 18:1 respectively and
no reduction in grass control was observed. The rate of flumetsulam in the prepackaged
mixture decreased as the metolachlor rate decreased but remained constant in the tank
mixture treatments. Our data indicate that flumetsulam and metolachlor were not additive
in the control of giant foxtail since grass control with flumetsulam tank mixtures was not
superior to the prepackaged mixture (Table 6, 8, and 10).

Chlorimuron, halosulfuron and flumetsulam alone provided up to 80 % control of
giant foxtail in field studies. Chlorimuron reduced grass number each year in soybean but
reduced dry weight per plant in only 1 of 3 years. In corn, halosulfuron alone reduced th e
number of giant foxtail in 1 of 3 years while weight per plant was reduced in all 3 years.
Reductions in grass dry weight were observed each year with flumetsulam treatments in
corn and in only 1 of 3 years in soybeans. The ALS inhibitors appear to retard early
season grass growth with control generally decreasing later in the growing season. The
earlier canopy cover and greater shading provided by com could assist the ALS inhibitors

in lengthier suppression of foxtail growth.

30

Neither additivity nor antagonism was consistently observed with any
ALS/metolachlor combination. In corn, plant dry weight was lower when an ALS inhibitor
was tank mixed with metolachlor compared to metolachlor alone in 1994. In 1995, a year
where grass control with metolachlor alone was good, flumetsulam, halosulfuron and
chlorimuron did not enhance or reduce giant foxtail control. In 1996 when grass control
with metolachlor alone was excellent, ALS-inhibiting herbicides did not decrease giant
foxtail control. Since these three ALS-inhibiting herbicides provide suppression of giant
foxtail, further experiments to determine the nature of metolachlor IALS inhibitor

interactions in grassy weed species are warranted.

31

Literature Cited

Herbicide Handbook, seventh edition. 1994. Pages 56-58, 131-133, 197-200, and 207-
209. Ahrens, W. H., ed. WSSA publication, Champaign, IL.

Kells, J. J. and K. A. Renner, et al. 1992. Annual Research Report, Michigan State
University. Pages 13-15. Michigan State University Publication, East Lansing MI.

Renner, K. A. and J. J. Kells, et al. 1996. Annual Research Report, Michigan State
University. Pages 13-15. Michigan State University Publication, East Lansing MI.

32

Table I. Metolachlor, flumetsulam, and halosulfuron plus furilazole treatments in
greenhouse studies.

 

 

Rate Rate Ratio
(kg ha“) (kg ha")
Metolachlor 0 - - -
0.14 - - ;
0.28 - - _
0.56 - - -
Metolachlor 0 flumetsulam 0.0073 -
0.14 flumetsulam 0.0073 19:1
0.28 flumetsulam 0.007 3 38: 1
0.56 flumetsulam 0.0073 77: 1
Metolachlor 0 flumetsulam 0.015 -
0.14 flumetsulam 0.015 10:1
0.28 flumetsulam 0.015 19:1
0.56 flumetsulam 0.015 38:1
Metolachlor 0 halosulfuron plus 0.011 -
furilazole
0.14 halosulfuron plus 0.011 13:1
fiirilazole
0.28 halosulfuron plus 0.011 27:1
furilazole
0.56 halosulfuron plus 0.011 53:1
furilazole
Metolachlor 0 halosulfuron plus 0.021 -
furilazole
0.14 halosulfuron plus 0.021 7 :1
furilazole
0.0.28 halosulfuron plus 0.021 13:1
furilazole
0.56 halosulfuron plus 0.021 27:1
furilazole

 

33

 

 

 

 

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43

Chapter 3
PHYSIOLOGICAL BASIS FOR THE ANTAGONISTIC RESPONSE OF GIANT
FOXTAIL (SE TARIA FABERI L.) TO METOLACHLOR IN THE PRESENCE OF
FLUMETSULAM, HALOSULFURON AND CHLORMURON-ETHYL.
Abstract. Combinations of metolachlor plus the ALS-inhibiting herbicides flumetsulam
and halosulfuron were less efficacious in controlling grasses compared to applications of
metolachlor alone. Greenhouse studies were conducted to quantify the activity of
metolachlor, flumetsulam, chlorimuron and halosulfuron with and without furilazole on
the growth of giant foxtail. Metolachlor at 0.28 kg ha" provided 97% control of giant
foxtail 28 days after treatment (DAT) as reflected in both plant height and dry weight
accumulation. Flumetsulam at 0.063 kg ha", chlorimuron at 0.031 kg ha“, and
halosulfuron with and without furilazole at 0.084 kg ha'1 reduced giant foxtail dry weight
70, 80, 40 and 63 % , respectively, 28 DAT. Antagonism of seedling shoot length occurred
only when the ALS inhibiting herbicides were applied 24 h prior to metolachlor, e. g. when
flumetsulam at 20 uM and chlorimuron at 50 uM were applied 24 h prior to metolachlor
at 5 “M and when flumetsulam at 50 ,uM was applied 24 h prior to metolachlor at 10 MM.
The radicle of giant foxtail emerged in 3 d while the shoot emerged between 2 to 11 d
later. This would allow a root absorbed herbicide to enter an emerging seedling prior to
a shoot absorbed herbicide. Site of uptake studies indicated that metolachlor and
chlorimuron were predominantly shoot absorbed by giant foxtail while flumetsulam and
halosulfuron with and without safener were root absorbed. Soil mobility studies indicated
that flumetsulam, halosulfuron and chlorimuron are more mobile than metolachlor on a

Capac sandy loam soil with 3.3% OM. Seeds treated with an ALS inhibitor 24 h prior to

metolachlor absorbed 2 to 5 times more metolachlor than seeds pretreated with
metolachlor, pretreated with metolachlor plus an ALS inhibitor or no pretreatment
(metolachlor alone). Giant foxtail converted over 80% of the l“C-metolachlor to metabolite
by 8 h after treatment when pretreated with an ALS inhibitor while only 11% was
metabolized in seeds exposed to metolachlor only. These data indicate that increased
metabolism of metolachlor as a result of exposure to an ALS inhibitor could result in

reduced grass control under field conditions.

INTRODUCTION

Metolachlor (2-chloro-N-(2-ethyl—6-methylphenyl)-N-(2-methoxy-l-methylethyl)
aetamide) is an acetanilide herbicide applied for annual grass control in corn (Zea mays)
and soybean (Glycine max) while flumetsulam (N-(2,6—difluorophentyl)-5-methyl
[1,2,4]triazolo [1 ,54]pyrimidine-2-sulfonamide) , chlorimuron (2-[[[[94—chloro—6-methoxy-
2-pyrimidinyl)amino]carbonyl] amino] sulfonyl]benzoic acid) and halosulfuron (methyl
3-chloro-5-(4,6-dimethoxypyrimidin-2-yl-carbamoylsulfamoyl)-1-methylpyrazole-4-
carboxylate) are three acetolactate synthase (ALS)l inhibiting herbicides that provide some
grass suppression (Anonymous 1993, Frear et al. 1993). In some field trials, combinations
of metolachlor plus flumetsulam or halosulfuron have resulted in reduced grass control
compared to control obtained from metolachlor alone (Kells and Renner 1992). This

interaction was also observed in other states under a variety of soil and environmental

 

‘Abbreviations: ALS, acetolactate synthase; DAT, days after treatment; DAPL,
days after planting; RH, relative humidity; Rf, ratio of front value; TLC, thin-layer
chromatography.

44

conditions (Owen et a1 1993, Simmons et al. 1993, Lueschen et al. 1995, Wax et a1.
1995). This antagonistic response was also observed under greenhouse conditions (Chapter
2). Factors that could influence the activity of preemergence herbicides included site of
herbicide uptake and soil mobility.

Metolachlor has been shown to be shoot absorbed by many grassy weed species
(Pillai et al. 1979) while the ALS inhibitors are root absorbed by most broad-leaved weed s
(Reddy and Bendixen 1989, Anonymous 1993). If the ALS inhibitors are absorbed
preferentially over metolachlor by giant foxtail (Setaria faberi Herrm.) as a result of
differences in site of uptake, then it is possible for the ALS inhibitors to interfere with
metolachlor efficacy.

Soil characteristics play an important role in any preemergence herbicide
interaction. Metolachlor has a water solubility of 530 ppm and a K0c =200 (Herbicide
Handbook 1994). The ALS inhibiting herbicides are weak acids and have water solubilities
and K0c values that are pH dependent. Both characteristics increase as soil pH increases
(Herbicide Handbook 1994). At a pH of 7, flumetsulam has a water solubility of 5600 ppm
and a Koc= 182, halosulfuron a solubility of 1630 ppm and Koc=94 and chlorimuron a
solubility of 1200 and K0,= 110. The higher water solubilities of the ALS-inhibiting
herbicides compared to metolachlor would indicate that a greater concentration of the AL S
inhibitors would be available for plant uptake at a higher soil pH.

The objectives of this research were to determine the conditions under which the
observed antagonism occurs and to determine the basis for the antagonism. Laboratory

experiments were conducted to confirm the antagonistic response of giant foxtail to

45

metolachlor in the presence of flumetsulam, chlorimuron and halosulfuron. If this
antagonistic response could be confirmed in the laboratory, experiments to determine the

physiological basis for the antagonism would then be initiated.

MATERIALS AND METHODS
Greenhouse experiments. Greenhouse experiments were conducted to quantify the activity
of metolachlor and ALS-inhibiting herbicides on giant foxtail growth. Giant foxtail seeds
were planted in 227 g pots containing a sandy loam soil (Capac sandy loam [Aerie
Ochraqualfs, fine loamy, mixed, mesic]; pH 6.8 with 3.3% organic matter). The soil field
capacity was determined to be 40% on a v/v basis and water was added to bring soil up
to field capacity. Herbicide treatments were applied to the pots using an 8003B nozzle at
209 L ha'1 at 207 kPa. Treatments included flumetsulam at 0.063, 0.031, 0.0063 and
0.0031 kg ai ha"; chlorimuron at 0.031, 0.016, 0.0031, and 0.0016 kg ai ha ;1 and
halosulfuron with and without furilazole [3-dichloro acetyl-5-(2-furanyl)-2,2-dimethyl-
oxazolidine] at 0.084, 0.042, 0.0084, and 0.0042 kg ai ha". Maximum herbicide rates
were based on field use rates for the ALS- inhibiting herbicides and ranged from 1 to 4
times the rates applied in previous greenhouse studies (Chapter 2). An untreated control
and metolachlor at 0.56 and 0.28 kg ai ha'1 were also included. All plots were uniformly
watered; alternating between surface watering and subirrigation each day. Pots were over
seeded and thinned to 10 grasses per pot 7 days after treatment (DAT) to prevent plant
competition. Pots within replications were re-randomized on a weekly basis. Plant height

(cm) and visual estimates of growth reduction were measured 21 and 28 DAT. Median

46

height was recorded to the nearest 0.5 cm. Visual evaluations compared the amount of
biomass in a pot to that of the untreated control. All above ground material was harvested
28 DAT, dried, and weighed to the nearest 0.1 mg on a per pot basis. The experiment
was conducted as a randomized complete block with four replications and repeated. An
ANOVA was completed on the data and the means separated by Fisher's protected LSD
at the 0.05 significance level. The data presented are the combined means of two
experiments containing four replications each.

Pregermination of giant foxtail seeds. All of the following experiments required giant
foxtail seeds to be pregerrninated with the exception of the emergence study.
Pregermination was conducted prior to initiation of the experiments. Giant foxtail seeds
were pregerrninated in a 0.06 M NaCl solution for 10 days. The salt solution was aerated
and changed daily. Preliminary studies showed a 12% decrease in variability of seedling
response if a 1 h imbibition was included prior to exposure to a herbicide (data not.
shown). Therefore, a 1 h distilled water imbibition period was included prior to exposure
of the pregerminated seeds to any herbicide.

Timing study. After pregermination, five seeds with not more than 2 mm of radical
emerged were placed on #1 Whatman2 filter paper in 100 by 20 mm petri plates. Seeds
were exposed to solutions of metolachlor at 5 and 10 uM, flumetsulam and halosulfuron
with and without furilazole at 10, 20 and 50 12M and chlorimuron at 50 11M. Seeds were
also exposed to combinations of metolachlor with these ALS-inhibiting herbicides. Each

treatment was applied with a fertilizer solution (one-half strength modified Hoagland’s

 

2 Whatman International Ltd. Clifton, NJ.

47

solution at pH 6.8) and without any fertilizer. A total of 7 ml of liquid per plate was added
initially with an additional 1 ml of water added 3 to 4 days later. Solution pH ranged from
6.5 to 7.1. Timing of herbicide introduction into the petri plates was varied to simulate
differences in seedling interception of metolachlor and the ALS-inhibiting herbicides.
Metolachlor was added 24 h prior to the ALS inhibitor, the ALS inhibitor added 24 h prio r
to metolachlor, and both herbicides were applied together. Seedlings were grown for 7
days in a growth chamber at 28 C day / 14 C night with a 16 h photoperiod. Root and
shoot lengths were measured 7 DAT. This experiment was a three factor factorial
(herbicide, rate, fertilizer) arranged in a randomized complete block design. The
experiment had five replications of each treatment and an untreated control for each
herbicide exposure timing. The experiment was repeated and means were combined over
the two experiments. Data presented are those means as separated by Fisher's protected
LSD at the 0.05 level of significance. Shoot length was more consistent in the untreated
controls and within a herbicide treatment so only shoot data will be presented.

Emergence study. Twenty giant foxtail seeds were placed in vials containing 20 ml of
0.06 M NaCl solution and allowed to grow for 14 days. Seeds were transferred to a
separate vial after emergence of the radicle structure so as to mark the progress of each
seed after emergence. Seedlings were transferred to a third vial after the emergence of the
shoot. Radicle and shoot emergence was determined by light microscopy and shoot and
root length was measured for each seed each day. This experiment contained twenty
replications and was repeated. Shoot and root data were compared using a Student’s t-test

analysis.

48

Site of uptake experiments. A charcoal barrier method was employed similar to that of
Pillai (Pillai 1979) to determine site of herbicide uptake. A l-cm layer of moist sandy loam
soil (Capac sandy loam; pH 6.8 with 3.3% organic matter) was placed in aluminum pie
tins, followed by a 1-cm layer of activated charcoal. This layer of charcoal was followed
by another l-cm layer of moist soil. Ten pregerrninated giant foxtail seedlings were placed
in the activated charcoal layer. Seedlings had no more than 2 mm of radicle exposed and
were completely isolated from the treated soil by the charcoal barrier. Herbicide treated
soil was placed alternately above or below the charcoal layer while the remaining soil layer
contained untreated soil. Preliminary greenhouse experiments determined the optimal
depth for giant foxtail emergence as 1 cm and also established herbicide application rates.
Metolachlor at 0.056 and 0.028 kg ha“, flumetsulam at 0.063 kg ha'1 , chlorimuron at
0.031 kg ha", and halosulfuron with and without safener at 0.084 kg ha" were sprayed
on the soil directly above or below the activated charcoal layer. A treated control
consisting of treated soil both above and below the charcoal layer and an untreated control
were also included. The study consisted of four replications of each treatment and was
repeated. Plants were grown in growth chambers with a 16 h photoperiod at 28 C day
/ 14 C night temperature regime with 90% relative humidity (RH). Pots were alternately
surface and subsurface watered every day. Shoot emergence, shoot height and root and
shoot dry weights was measured 28 days after planting (DAPL). Data was subjected to
ANOVA and treatment means separated using Fisher's protected LSD at the 0.05 level of
significance. Means presented were combined over two experiments. Trends with shoot

data were a more consistent indicator of plant response compared to root data. Therefore,

49

only the shoot data will be presented.
Soil mobility studies. Two laboratory experiments were conducted to determine the
relative soil mobility of metolachlor, flurnetsulam, chlorimuron and halosulfuron with and
without the safener furilazole. In the first experiment, soil thin-layer chromatography
(TLC) was used to assess the relative mobility of metolachlor, chlorimuron and
flumetsulam in a sandy loam soil (Capac sandy loam; pH 6.8 with 3.3 % organic matter).
Soil that had been screened to 50 mesh was layered to a 2-mm thickness on a 20 cm by 20
cm glass plate using a water slurry. A 18-ul droplet of l“C-metolachlor (uniformly ring
labeled; S.A.- 2900 kBq kg"), a 22-ul droplet of l“C-flumetsulam (uniformly aniline
labeled; S.A.- 800 kBq kg") and a 100-141 droplet of 1“C-chlorimuron (uniformly
pyrimidine labeled; S.A.- 1200 kBq kg") were spotted 3 cm from the bottom of the plate.
The quantity of each herbicide corresponded to 9.5 u g for all herbicides. Herbicide
solutions were spiked with approximately 5.0 x 10" dpm/aliquot of the appropriate
herbicide. The plates were divided into four to six lanes and developed in distilled water
to a distance of 15 cm. Plates were oven dried prior to quantification of radioactivity. The
ratio of front (R) values were measured and the radioactivity quantified using a plate
scanner. Data was then subjected to AN OVA and treatment means separated using Fisher's
protected LSD at the 0.05 level of significance.

Soil column mobility experiments were then conducted to compare the relative
mobility of flumetsulam and chlorimuron to halosulfuron with and without furilazole.
Plastic tubes measuring 36 cm in length with an internal diameter of 5 cm were vertically

cut and sealed with duct tape. The columns were then filled with a sandy loam soil (Capac

50

sandy loam; pH 6.8 with 3.3 % organic matter) with 10 cm of space left at the top of each
tube. Cheesecloth was used to seal the bottom of the columns. The soil in the columns
was brought up to field capacity (40% moisture v/v) through surface watering and allowed
to sit overnight. After the tubes were conditioned for 24 h, metolachlor, flumetsulam,
chlorimuron or halosulfuron with and without safener were added. Metolachlor rates were
0.56 and 1.12 kg ha" and rates for the ALS-inhibiting herbicides corresponded to the field
rates used in the site of uptake experiments. Each column received water to the equivalent
of 2.5 cm of rain directly after herbicide application and allowed to elute overnight.
Columns were then cut open vertically and redroot pigweed (Amaranthus retroflexus L.)
planted as an indicator species. Redroot pigweed was used as an indicator since all of the
herbicides control this weed (Kells and Renner 1997). Herbicide movement was
determined by redroot pigweed response relative to that of the untreated control 7 DAT.
Since the sensitivity of pigweed to metolachlor is not known compared to the ALS
inhibitors, metolachlor mobility relative to the ALS inhibitors can not be stated with
certainty from this study. Distance (cm) was measured from the top of the soil to the first
unaffected plant. This was considered to be the distance moved by the herbicide. There
were four columns per herbicide treatment and each half of the column was planted, such
that there were two samples in each of the four replications. This study was repeated and
means for the two experiments were combined and separated using ANOVA. Means were
then subjected to Fisher’s protected LSD at the 0.05 level of significance.

Absorption study. Giant foxtail seedlings were pretreated for 24 h with 10 11M

metolachlor, 50 uM flumetsulam, 50 uM chlorimuron, 50 HM halosulfuron with and

51

without safener or a combination of metolachlor plus these ALS-inhibiting herbicides.
After pretreatment, seeds were placed in test tubes and exposed to l4C-metolachlor
(uniformly ring labeled; S.A.- 2900 kBq kg") plus a non-labeled ALS-inhibiting herbicide.
The experiment also included plants exposed only to l"C-metolachlor (not pretreated) and
an untreated control (Table 7). The test tubes were then placed on a shaker until sampling.
Ten seedlings per treatment were sampled at 0, 0.5, 1, 2, 4 and 8 h after exposure to 14C-
metolachlor with four replications at each sample time. The seedlings were triple rinsed
with methanol prior to combustion in a Harvey OX3003 biological oxidizer and liquid
scintillation4 was used to determine the total amount of 1‘C absorbed. Data was then
subjected to mean separation using Fisher’s protected LSD at the 0.05 level of
significance.

Metabolism study. This experiment was conducted as a pulse—chase experiment utilizing
”C-labeled metolachlor (uniformly ring labeled; S.A.- 2900 kBq kg"). All other
herbicides and safeners were not radioactive. Grass seedlings were placed in a solution
containing 10 MM metolachlor alone, 10 uM of metolachlor plus 50 MW of either
flumetsulam, chlorimuron or halosulfuron (with and without furilazole), the ALS-
inhibiting herbicides alone, or distilled water. After this 24 h pretreatment, the seeds were
transferred to fresh solutions that contained 10 12M l‘C-metolachlor plus 50 MM of an
ALS-inhibiting herbicide or 10 uM l‘C-metolachlor only. After 1 h, seeds were rinsed for

1 min with distilled water, and placed into a vial containing a solution of the

 

3 R. J. Harvey Instrument Corp. Patterson, NJ 07642.
‘ Model 1500. Packard Instrument Corp. Downers Grove, IL 60515.
52

corresponding herbicide combination substituting unlabeled metolachlor for 1"C-
metolachlor. Vials were then placed on a shaker and vials removed at 0, 0.5, 1, 2, 4, and
8 h after pulsing. Plant samples were rinsed with methanol for l min and stored at -30 C
until extraction. The extraction procedure used was similar to that of Rowe et al. (1990)
for metolachlor extraction from corn. A 0.26 g sample (approximately 100 seeds) per
treatment was extracted with a grinder in 100% methanol for 3 min. The extraction
efficiency was greater than 92 % for both repetitions of the experiment. The extract was
reduced under a vacuum and 1 ml of 90% methanol was added for reconstitution. A 100
ttl aliquot of extract for each treatment and unextracted metolachlor were spotted on silica
gel TLC plates5 previously divided into lanes. The plates we re developed using a mixture
of butanol:acetic acidzwater (12:3 :5 v/v). There were four replications of each treatment
and the experiment was repeated. R, values were calculated from TLC plates
measurements and radioactivity quantified using a plate scanner. The TLC plates were
evaluated for quantitative and qualitative changes in the C 1‘—metabolites in the presence and
absence of an ALS inhibitor. Treatment means were separated using Fisher’s protected

LSD at P=0.01 after t-test analysis.

RESULTS AND DISCUSSION
Greenhouse experiment. Metolachlor alone at 0.56 and 0.28 kg ha" reduced shoot height

of giant foxtail 99 and 97 % , respectively, 28 DAT (Table 1). Giant foxtail shoot heights

 

5 Whatman Silica Gel 150A LK5DF, Whatman International Ltd. Maidstone,
UK.

53

and shoot dry weights (Table l) were reduced by the ALS-inhibiting herbicides at the field
use rates. Chlorimuron at 0.031 kg ha'1 reduced giant foxtail height by 63% while
flumetsulam at 0.063 kg ha" only reduced grass height by 44% compared to the untreated
control. Halosulfuron alone at 0.084 kg ha" decreased grass height by 31% while
halosulfuron plus furilazole decreased grass height by 60% (Table 1). This increase in
grass control when the safener furilazole is added to halosulfuron implies that furilazole
has herbicidal activity with respect to giant foxtail. These results contradict earlier find ing
that stated furilazole should act as a safener against acetanilide herbicides (Bussler et al
1991). The herbicidal activity of furilazole could be due to the similarity in structure
between the safener and the acetanilide herbicides. Plant dry weight data 28 DAT showed
a similar trend as that observed from plant height measurements (Table 1). These data
rank the giant foxtail activity of the ALS inhibiting herbicides as chlorimuron >
halosulfuron plus furilazole > flumetsulam > halosulfuron at the field use rate.

Timing study. Fertilizer regime did not influence the response of giant foxtail to
metolachlor, flumetsulam, or chlorimuron, therefore, data was combined over fertilizer
regimes. Antagonism of giant foxtail control with metolachlor plus flumetsulam
combinations was observed only when flumetsulam was applied 24 h prior to metolachlor.
Metolachlor at 5 uM plus flumetsulam at 20 uM and metolachlor at 10 ,uM plus
flumetsulam at 50 “M resulted in increased shoot length compared to metolachlor (Table
2). Neither an increase nor decrease in giant foxtail control was noted from other
metolachlor plus flumetsulam combination compared to metolachlor alone, regardless of

application timing. Metolachlor at 5 HM and chlorimuron at 50 11M resulted in increased

54

shoot length compared to metolachlor alone when chlorimuron was applied prior to
metolachlor (Table 2). This increase in shoot length was not observed when the
metolachlor rate was increased to 10 ,uM. All other combinations did not increase or
decrease shoot length compared to metolachlor alone.

Shoot length with halosulfuron decreased upon the addition of fertilizer to the
system (Table 3). Conversely, shoot lengths with halosulfuron plus furilazole were
increased upon the addition of fertilizer (Table 3). The concentration of Ca + * has been
linked to the opening and closing of ion channels in plant roots (Wilkinson and Duncan
1993). This effect was partially responsible for differences in ion absorption mechanisms
as well as plant responses to stress. The increase in shoot length could be a result of
furilazole safening the giant foxtail seedling against absorption of an ion that could cause
a detrimental effect in the presence of halosulfuron.

All incidences of increased shoot growth when halosulfuron was applied with
metolachlor occurred in the presence of fertilizer and when halosulfuron was applied first.
Metolachlor at 10 uM plus halosulfuron at 10 or 50 MM and metolachlor at 5 ttM plus
halosulfuron at 20 [1M resulted in antagonism compared to metolachlor alone (Table 3).

Combinations of metolachlor at 10 MM and halosulfuron plus furilazole at 10
uM, metolachlor at 5 “M and halosulfuron plus furilazole at 20 MM in the presence of
fertilizer and metolachlor at 10 uM and halosulfuron plus furilazole at 50 uM in the
absence of fertilizer resulted in increased shoot growth when the ALS inhibitor was added
first (Table 3).

In summary, giant foxtail shoot length was not increased compared to metolachlor

55

alone when ALS inhibitors were applied with or 24 h after metolachlor. Incidences of
herbicide antagonism occurred only when the ALS inhibiting herbicides were applied 24
hours prior to metolachlor. This occurred when metolachlor at 5 uM was applied after
flumetsulam at 20 uM and chlorimuron at 50 HM and when metolachlor at 10 uM was
applied after flumetsulam at 50 ttM. Combinations of metolachlor and halosulfuron with
and without furilazole was influence by fertilizer regime. When an ALS-inhibiting
herbicide was applied first, reduced grass control was observed in 25% of the
halosulfuron, 33 % of the flumetsulam and l of 2 chlorimuron treatment combinations.
Emergence study. Germination of giant foxtail ranged from 25 to 30 %. In every instance ,
germination of giant foxtail started with the emergence of a radicle followed later by
emergence of the shoot. Radicle emergence occurred an 2 to 11 (1 prior to shoot emergence
(Figure l). Seedling abortion following radicle emergence was rare but did occur on
occasion (less than 0.5 % of the total germination). While some precocious germinating
seeds were present, they comprised less than 2% of the overall seedlings. This temporal
difference in structural emergence could be a means by which a root absorbed ALS-
inhibiting herbicide could enter the plant prior to a shoot absorbed herbicide.

Site of uptake experiments. Differences were observed in the site of uptake for
metolachlor and the ALS inhibitors. Metolachlor entered predominantly through the
emerging shoot of giant foxtail (Table 4). Flumetsulam and halosulfuron both with and
without furilazole entered through the radicle with essentially no shoot absorption.
Conversely, the primary site of uptake for chlorimuron in giant foxtail was the shoot. A

decrease in growth reduction was noted when comparing the shoot absorption data to the

56

treated control for chlorimuron (Table 4). Theoretically, the growth reduction from
chlorimuron above the charcoal barrier should not be significantly different from
chlorimuron applied both above and below the barrier if the site of uptake was the shoot
(Pillai et al. 1979).The increase in shoot length and shoot dry weight can not be explained.
Differences in the site of uptake between metolachlor compared to flumetsulam and
halosulfuron may be a factor in the antagonism of giant foxtail control. Since chlorimuro n
and metolachlor have the same site of uptake, another explanation is necessary for the
interaction between these herbicides.

Soil mobility study. Soil TLC studies showed the order of mobility of the herbicides to
be flumetsulam > chlorimuron > metolachlor. The R, value for metolachlor was 0.34
compared to 0.98 and 0.88 for flumetsulam and chlorimuron, respectively (Table 5).
Flumetsulam R, values have not been published but studies have shown chlorimuron soil
Rf values to be 0.8 for sandy soil with lower organic matter and pH (Goetz 1989). Radio
scans of the TLC plates show streaking in lanes where flumetsulam and chlorimuron were
applied indicating sorption of these compounds to soil colloids. Regardless of herbicide
sorption, less than 16 % of the flumetsulam or 31% of the chlorimuron applied remained
at the origin while 65 % of the metolachlor was detected at the origin (Table 5).

Relative mobility of these ALS-inhibiting herbicides determined in soil TLC
experiments was comparable to studies conducted in soil columns (Table 6). The order of
mobility as determined by soil columns was flumetsulam > chlorimuron > halosulfuron with
furilazole =halosulfuron. Since the ALS inhibitors were more mobile in the soil compared

to metolachlor, there is the potential for an emerging giant foxtail seedling to absorb an

57

ALS inhibitor prior to metolachlor. The greater mobility of the ALS inhibitors in concert
with a greater ALS inhibitor concentration increases the chances of ALS interception prior
to metolachlor, thereby, increasing the probability of an interaction occurring.
Absorption study. Total metolachlor absorption by giant foxtail ranged from 2 to 18 %
8 h after treatment (Table 7). Pretreatment with metolachlor resulted in only 2%
absorption of l"C-metolachlor, regardless of the ALS-inhibiting herbicides. Therefore, data
for these pretreatments were combined over the ALS inhibitors in Figure 3. Absorption
patterns between plants pretreated with metolachlor and plants pretreated with an ALS
inhibitor plus metolachlor were similar at sample times greater than or equal to 2 h
(Figures 2, 3). Seeds absorbed twice the amount of metolachlor 8 h after treatment when
pretreated with flumetsulam plus metolachlor as compared to chlorimuron plus metolachlor
or halosulfuron (with and without furilazole) plus metolachlor pretreatments (Figure 3).
Seedlings never exposed to an ALS inhibitor absorbed more I‘C-metolachlor compared to
metolachlor pretreatments and ALS inhibitor plus metolachlorpretreatments (Figures 2,
3). Absorption took place at a more rapid rate when there was no ALS inhibitor present
with significant absorption occurring at 2 h compared to 4 h after treatment for the
metolachlor and metolachlor plus an ALS inhibitor pretreatments.

Seedlings receiving a pretreatment of an ALS inhibitor absorbed more metolachlor
compared to the other pretreatments (Figures 3). This was unexpected since previous data
showed a decrease in metolachlor efficacy when an ALS-inhibiting herbicide entered the
seedling first. Plants pretreated with chlorimuron absorbed approximately 19 % of the total

l4C-metolachlor 8 h after treatment compared to flumetsulam which absorbed 16 % and

58

halosulfuron alone or halosulfuron with furilazole absorbed which absorbed a total 13 %
and 12% 8 h after treatment, respectively (Figure 3). The induction of a metolachlor
detoxification pathway could explain the increase in uptake but decrease in grass control
observed when a grass seed is exposed to an ALS inhibitor prior to metolachlor.

These data would also imply that metolachlor is taken up in a passive manner since
metabolism and thus removal of metolachlor from the site of uptake would cause more
metolachlor to be taken up by giant foxtail. Cottingham and Hatzios (1992) found that
susceptible varieties of corn absorbed more metolachlor than tolerant varieties. Ashton and
Monaco (1991) state that translocation of metolachlor was predominantly via the apoplast
in grass species implying metolachlor is entering the plant through passive uptake. Our
data support passive uptake as the method of entry of metolachlor in giant foxtail.
Metabolism study. R, values for the parent metolachlor and the metabolite, assumed to
be the glutathione conjugate of metolachlor, were 0.82 and 0.49, respectively. This data
is consistent with the R, values reported by Rowe et al (1990) for tolerant versus
susceptible corn species.

Twice the amount of metolachlor was absorbed during the l h pulse period when
seeds were pretreated with an ALS inhibitor compared to pretreatments of metolachlor
and metolachlor plus ALS-inhibiting herbicide combinations (Figures 4, 5). The re was no
significant difference in the amount of metolachlor absorption among the other
pretreatments.

Giant foxtail is a susceptible species so the percent of metolachlor metabolized was

low (Figures 4, 5). Initial metabolism was minimal with detectable metabolite formation

59

1 h after treatment for ALS pretreated seeds, 2 h after treatment for seeds pretreated with
metolachlor plus ALS inhibitor combinations, and 4 h after treatment for seeds pretreated
with metolachlor or treated with metolachlor alone (Figures 4, 5). No significant
differences between ALS-inhibiting herbicides were observed when seeds were pretreated
with metolachlor or when seeds were pretreated with metolachlor plus an ALS inhibitor
at the P=0.01. Therefore, the data were combined over the ALS inhibitors for these
pretreatments (Figure 4). Metabolism 4 and 8 h after treatment was significantly greater
in seeds pretreated with metolachlor plus an ALS inhibitor compared to seeds that were
pretreated with metolachlor or exposed to metolachlor only. In the treatment where seeds
were exposed to metolachlor in the absence of an ALS inhibitor, metabolism was lower
than any other treatment. This indicates that any exposure to an ALS inhibitor increases
the metabolism of metolachlor regardless of exposure timing.

There was a significant difference between seeds pretreated with an ALS inhibitor
and other seed pretreatments (Figure 5). Metabolism of metolachlor in seedlings pretreated
with an ALS inhibitor was more rapid than other pretreatments and a greater percentage
of parent was converted to metabolite during the course of the experiment. There was no
difference between ALS inhibitor pretreatments 8 h after treatment, though the
halosulfuron pretreatment showed significantly less metabolism 2 and 4 h after treatment
compared to the other ALS inhibitors. Giant foxtail converted over 80% of the 1“C-
metolachlor to metabolite 8 h after treatment when pretreated with any ALS inhibitor while
only 11% was converted in seeds exposed to metolachlor in the absence of an ALS

inhibitor (Figure 5). This data indicate that increased metabolism of metolachlor, as a

60

result of exposure to an ALS inhibitor, could be the basis for the antagonism sometimes
observed in the field.

These data would indicate that there is an induction of a metabolic pathway for
metolachlor when a giant foxtail seedling intercepts an ALS inhibiting herbicide regardless
of the timing of seedling interception. The greatest impact of this increase in metolachlor
breakdown occurs when the giant foxtail seed intercepts the ALS inhibitor 24 h prior to
metolachlor. Since antagonism of grass control could only be observed when seeds were
pretreated for 24 h with an ALS inhibitor (Table 3) and not with any other pretreatment,
the antagonism observed in the preliminary field trials may be the result of an induction
of more rapid and greater overall metolachlor metabolism.

A stimulation of glutathione production as well as increased glutathione-S-
transferase activity have been linked to increased metolachlor metabolism (Dean et al.
1990, Cottingham and Hatzios 1992). The cytochrome P450 enzyme integral in the
breakdown of the ALS inhibitors (Frear et al. 1993) has also been implicated as a route
of metolachlor degradation in plants (Cole and Owen 1987, Herbicide Handbook 1994,
Walton and Casida 1995). Conversion of metolachlor to its glutathione congugate takes
place through hydroxylation and dechlorination to an oxamic acid which is known to
involve both GSH/GST and P450 pathways (Miaullis et al. 1978). Herbicides have been
shown to protect plants from subsequent exposure of similar herbicides through an
initiation of detoxification pathways (Ezra et al.1985) but not from different classes of
herbicides. It can be hypothesized that the ALS inhibitors can induce P450 activity in such

a way as to stimulate metolachlor metabolism through enhancement of GSH production

61

or GST activity.

Future studies of interest would involve confirming increased metabolism in other
susceptible annual grass species such as barnyardgrass. Also of interest is the metabolism
of metolachlor in corn and soybean. This would determine if the ALS inhibitors increase
the rate of metolachlor metabolism in tolerant species as well. In addition, quantitation of
GSH production and GST activity in giant foxtail when exposed to the various
pretreatments could elucidate the exact mechanism by which increased metolachlor
metabolism occurs.

A final area of interest would be investigating the possibility of using an ALS
inhibitor as a safener for grassy crops. Graminaceous crops, such as sorghum or wheat,
could be safened against metolachlor injury if an ALS inhibitor was applied first. ALS
inhibitors labeled, or soon to be labeled, for use in these crops could be tested for potential

safening against metolachlor.

62

Literature Cited

Anonymous. 1993. Mon 12000 technical data sheet. Monsanto publication, Monsanto
Agric. Co. MAC-12000—01. PP. 1-4.

Anonymous. 1993. Mon 13900 technical data sheet. Monsanto publication, Monsanto
Agric. Co. MAC-13900-01. PP. 1-2.

Ashton, F. M. and T. J. Monaco. 1991. Aliphatics, amines ans amino acids, p 155. in
Weed Science: Principals and Techniques, 3rd edition. Wiley Interscience.

Bussler, B. H., R. H. White, and E. L. Williams. 1991. Mon 13900: a new safener for
gramineous crops. Brighton Crop Protection Conference. Weeds pp. 39-44.

Cole, D. J ., and W. J. Owen. 1987. Influence of monooxygenase inhibitors on the
metabolism of the herbicides chlortoluron and metolachlor in cell suspension cultures.
Plant Sci. 50: 13-20.

Cottingham, C. K. and K. K. Hatzios. 1992. Basis of differential tolerance of two corn
hybrids (Zea mays) to metolachlor. Weed Sci. 40:359-363.

Dean, J. V., J. W. Gronwald And C. V. Eberlein. 1990.1nduction of glutathione S-
transferase isozymes in sorghum by herbicide antidotes. Plant Physiol.92:467-473.

Ezra, G., D. G. Rusness, G. L. Lamoureux and G. R. Stevenson. 1985. The effect of

CDAA (N, N—diallyl-2-chloroacetamide) pretreatments on subsequent CDAA injury to corn
(Zea mays). Pestic. Biochem. Physiol. 23: 108-115.

Frear, D. S., H. R. Swanson, and F. S. Tanaka. 1993. Metabolism of flumetsulam (DE-
498) in wheat, corn and barley. Pestic. Biochem. Physiol. 45: 178-192.

Goetz, A. J ., R. H. Walker, G. Wehtje, and B. F. Hajek. 1989. Sorption and mobility of
chlorimuron in Alabama soils. Weed Sci. 37:428-433.

Herbicide Handbook. 1994. “Chlorimuron” ,“Flumetsulam” , “Metolachlor”, “MON
12000". Weed Sci. Soc. Amer. Publication pp. 56-58, 131-133, 197-200, 207-209.

Kells, J. J. and K. A. Renner. 1997. Weed Control Guide for field crops. Michigan State
University Research Extension Bulletin E-434 pp.62, 106.

Kells, J. J. and K. A. Renner, et al. 1992 Weed Control Results in Field Crops. Michigan
State University Research Report pp. 13-15.

63

Lueschen, W. E. and J . K. Getting. 1995. Broadleaf weed control in corn with soil
applied and postemergence herbicides at Lamberton, MN in 1995. N.C.W.S.S. Annual
Research Report. N.C.W.S.S. publication. 52: 220-221.

Miaullis, J. B., V. M. Thomas, R. A. Gray, J. J. Murphy and R. M. Hollingsworth.
1978. Metabolism of R-257 88 (N, N-diallyl-Z—chloroacetamide) in corn plants, rats and
soil. in Chemistry and Action of Herbicide Antidotes. F. M. Pallos, J. E. Casida, eds.
Academic Press, New York, pp 109-131.

Owen, M. J ., J. F. Lux and K. T. Pecinovsky. 1993. Evaluation of flumetsulam plus
metolachlor prepackage, metribuzin and other herbicides for weed management in no-
tillage systems, Ames, IA, 1993 N.C.W.S.S. Annual Research Report. N.C.W.S.S.
publication. 50: 306-307.

Pillai, P., D. E. Davis and B. Truelove. 1979. Effects of metolachlor on germination,
growth, leucine uptake and protein synthesis. Weed Sci. 27:634-637.

Reddy, K. N. and L. E. Bendixen. 1989. Toxicity, absorption and translocation of soil
applied chlorimuron in yellow and purple nutsedge (cyperus esculentus and C. rotundus).
Weed Sci. 37:147-151.

Rowe, L., E. Rossman and D. Penner. 1990. Differential response of corn hybrids and
inbreds to metolachlor. Weed Sci. 38:563-566.

Simmons, F. W., L. M. Wax and D. J. Maxwell. 1993.Evaluation of imazethapyr and
other herbicides for preemergence and postemergence weed control in imazethapyr
resistant corn, Kilbourne, IL, 1993. N.C.W.S.S. Annual Research Report. 50:202-203.

Walton, J. D. and J. E. Casida. 1995. Specific binding of an dichloroa cetamide herbicide
safener in maize at a site that also binds thiocarbamate and chloracetanilide herbicides.
Plant Physiol. 109:213-219.

Wax, L. M., S. E. Hart and D. J. Maxwell. 1995. Weed control systems for no-till
soybeans. Urbana, IL, 1995. N.C.W.S.S. Annual Research Report. 52:408-409.

Wilkinson, R. E. and R. R. Duncan. 1993. Calcium (“Ca“) uptake in GP-lO sorghum
root tips as influenced by hydrogen ion (H *) concentration and hours of exposure to H +-
ATPase inhibitors. J. Plant Nutr. 16(4):643—652.

Table 1. Reduction in giant foxtail shoot height and dry weight by metolachlor,
chlorimuron, flumetsulam and halosulfuron with and without furilazole 28 days after
treatment.

 

 

 

Herbicide Rate Shoot height Dry weight reduction‘l
reduction
(kg“ ha) (%)b (%)
Chlorimuron 0.031 63 80
0.016 46 59
0.0031 18 +14
0.0016 21 0
Flumetsulam 0.063 44 70
0.031 38 40
0.0063 3 1 55
0.0031 22 25
Halosulfuron 0.084 3 1 40
0.042 3 1 47
0.0084 9 24
0.0042 3 + 12
Halosulfuron plus 0.084 60 63
furilazole
0.042 36 54
0.0084 15 30
0.0042 18 35
Metolachlor 0.56 99 94
0.28 97 97
LSD,» 12 16

 

“Giant foxtail in the untreated control weighed 3.2 g/ 10 plants.
b % growth reduction values preceded by a ‘ + ’ indicate growth enhancement compared to
the untreated control.

65

Table 2. Giant foxtail shoot length as influenced by metolachlor combinations with
flumetsulam and chlorimuron when flumetsulam and chlorimuron were applied 24 h prior
to metolachlor. An asterisk denotes antagonism as determined by Colby’s multiplicative
model‘.

 

 

Rate Metolachlor
ALS Inhibitor
5 aM 10 “M
(HM) --- ------ (% of control)-------
Flumetsulam 10 1 1 12
20 17* 9
50 12 23*
Chlorimuron 50 16* 1 1

 

‘Untreated shoot length was 35 cm.

66

Table 3. Giant foxtail shoot length as influenced by metolachlor combinations with
halosulfuron with and without furilazole in the presence and absence of fertilizer when
halosulfuron was applied 24 h prior to metolachlor“.

 

 

ALS Rate Metolachlor
inhibitor
5 12M 10 HM
(,uM) with fertb no fertc with fert no fert
Halosulfuron 10 17 9 22* 7
20 23 * l4 6 1 1
50 3 1 1 26* 1 1
Halosulfuron 10 4 15 17* 8
+ furilazole 20 20* 7 7 9
50 6 10 15 31*

 

‘Untreated shoot length for fertilized control was 33 cm and for unfertilized control was
35 cm.

bwith fert indicates the presence of V2 strength nutrient solution in the system.
°no fert indicates the absence of nutrient solution in the system.

67

Table 4. Site of uptake in giant foxtail for metolachlor, flumetsulam, chlorimuron and
halosulfuron with and without furilazole.

 

 

 

 

 

 

Growth reductiona
Herbicide Rate Soil
placement Shoot length Grass dry weight
(kg" ha) (%)
Chlorimuron 0.031 Above 55 63
Below 12 27
Both 43 45
Flumetsulam 0.063 Above 9 14
Below 45 45
Both 37 41
Halosulfuron 0.084 Above 17 22
Below 54 49
Both 50 52
Halosulfuron 0.084 Above 9 16
+ furilazole Below 33 32
Both 28 31
Metolachlor 0.056 Above 52 55
Below 21 26
Both 46 49
0.028 Above 37 36
Below 14 18
Both 41 40
LSD,” 9 13

 

‘Giant foxtail shoot length and dry weight for the untreated control was 52 cm and 0.1 g
per plant for the respective measurements.

68

Table 5. Relative soil mobilities of metolachlor, flumetsulam and chlorimuron as compared
on soil thin-layer chromotography plates.

 

Herbicide Distance moved R, Herbicide Total Herbicide
value at origin herbicide remaining

 

 

at origin

------- (cm)------- (CPM) (CPM) ---- (%)-«-
Chlorimuron 13.2 0.88 6268 20218 31
Flumetsulam 14.8 0.98 3175 19850 16
Metolachlor 5.1 0.34 5195 15892 67
LSD,” 10

 

69

Table 6. Relative soil mobilities of metolachlor, flumetsulam, chlorimuron and
halosulfuron with and without the safener furilazole as compared on soil columns. The
indicator species used was redroot pigweed which was solid seeded down the length of the
tube.

 

 

 

Herbicide Rate Distance moved

0%" ha) (cm)

Chlorimuron 0.031 17.1
Flumetsulam 0.063 21.2
Halosulfuron 0.084 12.8
Halosufuron+furilazole 0.084 13.2
Metolachlor 1 . 12 5 .3
0.56 5.1

LSD,” 0.5

 

7O

Table 7. Giant foxtail absorption of metolachlor after 8 h after pretreatments of
metolachlor, an ALS inhibitor and a combination of metolachlor plus an ALS inhibitor.

 

 

 

Pretreatment Treatment 1"C-metolachlor
8 h after
treatment
( % absorbed)

Metolachlor 1“C-metolachlor+flumetsulam 2

Metolachlor l"C-metolachlor+chlorimuron 2

Metolachlor “C-metolachlor+halosulfuron 2

Metolachlor l"C-metolachlor+halosulfuron+safener 2

Flumetsulam l4C-metolachlor+flumetsulam 16

Chlorimuron l"C-metolachlor+chlorimuron 18

Halosulfuron 1"C-metolachlor+halosulfuron 13

Halosulfuron l"C-metolachlor+halosulfuron-l-safener 12

+ safener

Metolachlor+ 1"C-metolachlor-l-flumetsulam 7

flumetsulam

Metolachlor+ l"C-metolachlor+chlorimuron 3

chlorimuron

Metolachlor+ 1"C-metolachlor+halosulfuron 3

halosulfuron

Metolachlor+ l"C-metolachlor+halosulfuron-l-safener 4

halosulfuron +

safener

None Metolachlor 9

LSDm O. 85

 

71

Figure 1. Radicle and shoot emergence of giant foxtail over time.

72

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73

Figure 2. Absorption of metolachlor by giant foxtail as influenced by ALS-inhibiting
herbicides plus metolachlor pretreatments. Equation for the lines: lny =a+be"‘.
Metolachlor only r2=0.98; metolachlor pretreatment (combined over ALS inhibitors)
r2=0.99; chlorimuron plus metolachlor r2=0.90; flumetsulam plusmetolachlor r2=0.72;
halosulfuron plus metolachlor r2=0.84; halosulfuron plus furilazole plus metolachlor
r2 =0.86.

74

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75

Figure 3. Absorption of metolachlor by giant foxtail as influenced by ALS-inhibiting
herbicide pretreatments. Equation for the lines: lny =a+be"‘. Metolachlor only r2=0.98;
metolachlor pretreatment (combined over ALS inhibitors) r2 =0.99; chlorimuron r2 =0.98;
flumetsulam r2=0.94; halosulfuron r2=0.94; halosulfuron plus furilazole r2=0.94.

76

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77

Figure 4. Metabolism of metolachlor by giant foxtail as influenced by ALS-inhibiting
herbicides plus metolachlor pretreatments. Equations for the lines: metolachlor only y'
1=a+bx3 r2=0.63; metolachlor pretreatment (combined over ALS inhibitors) y'
1=a+bx21nx r2=0.45; ALS plus metolachlor (combined over ALS inhibitor) y‘l =a+bx2
r2 = 0.91.

78

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79

Figure 5. Metabolism of metolachlor by giant foxtail as influenced by ALS-inhibiting
herbicides; chlorimuron r2=0.99; flumetsulam r2=0.94; halosulfuron r2=0.99;
halosulfuron +furilazole r2=0.99

80

 

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82

Chapter 4

SUMMARY AND CONCLUSIONS

Antagonism of giant foxtail control was documented in preliminary field trials in
1992 and 1993 when metolachlor plus flumetsulam or halosulfuron was applied c ompared
to metolachlor alone. This antagonism could not be linked to edaphic or environmental
factors nor could this interaction be predicted with any certainty.

Greenhouse studies confirmed the presence of an interaction between metolachlor
and the ALS inhibitors. The frequency of antagonism increased as the ratio of metolachlo r
to ALS inhibitor decreased. There was no consistent increase or decrease in annual grass
control when metolachlor was applied with flumetsulam, halosulfuron or chlorimuron
compared to metolachlor alone in three years of field trials in corn and soybean. When
metolachlor activity was low, the ALS inhibitors should have increased grass control
compared to metolachlor alone since these ALS-inhibiting herbicides are more soluble than
metolachlor and they suppressed grass dry weight in three of three years in corn and
soybean.

The activity of the ALS inhibitors on giant foxtail was quantified and the order of
activity was chlorimuron > halosulfuron plus furilazole > flumetsulam > halosulfuron.
Experiments were then conducted in the laboratory to determine if antagonism was related
to the timing of seedling exposure to the herbicides. It was determined that antagonism
occurred if a giant foxtail seedling was exposed an ALS-inhibiting herbicide 24 h prior to
exposure to metolachlor but not with other exposure timings. The ratios of me tolachlor to

the ALS inhibitor used in these laboratory experiments were significantly less (1 :5, 1:4,

1:2, 1:1) than field applied rates (38:1 for metolachlorzflumetsulam prepackaged
mixtures). This data supported our previous research linking the frequency of antagonism
to ALS inhibitor concentration.

Timing of seedling interception of metolachlor was shown to play an integral role
in the expression of antagonism of giant foxtail control. An ALS inhibitor could be
intercepted prior to metolachlor through differences in site of uptake and/or soil mobility.
Emergence studies were performed to determine the timing of structural emergence for
giant foxtail. The radicle of giant foxtail emerged 3 d after experimental initiation while
shoot emergence occurred between 5 and 14 d. This temporal difference would allow a
root absorbed herbicide to be taken up by giant foxtail prior to a shoot absorbed herbicide.
Metolachlor and chlorimuron were shown to be predominantly shoot absorbed while
flumetsulam and halosulfuron (with and without furilazole) were root absorbed.
Differences in site of uptake could result in flumetsulam or halosulfuron interception prior
to metolachlor though another explanation was necessary for chlorimuron. Soil mobility
studies showed the ALS inhibitors were more mobile in the soil compared to metolachlor.
The ALS inhibitors, therefore, have the potential to encounter an emerging giant foxtail
seedling prior to metolachlor especially under low moisture, high pH conditions. This
greater mobility in concert with the greater solubility of the ALS inhibitors increases the
chance of ALS interception prior to metolachlor, thereby, increasing the probability of an
interaction occurring.

Absorption of metolachlor was increased when giant foxtail seedlings were
pretreated with an ALS inhibitor compared to other pretreatments. This increase in

absorption in concert with the observed decrease in herbicidal activity suggest passive

83

uptake of metolachlor and an induction of metolachlor metabolism. Metolachlor
metabolism was increased in the presence of an ALS inhibitor compared to metolachlor
alone regardless pretreatment. When seedlings were pretreated with an ALS inhibitor,
more rapid and more complete metabolism of metolachlor was observed. These data also
indicate an induction of a metabolic pathway for metolachlor when a giant foxtail seedling
intercepts an ALS inhibiting herbicide regardless of when the seed intercepts these
herbicides. The greatest impact of this increased metabolism occurred when the giant
foxtail seedling intercepted the ALS inhibitor 24 h prior to metolachlor. Since antagonism
of grass control could only be observed when seeds were pretreated with an ALS inhibito r
and not with any other pretreatment, the antagonism observed in the field appears to be
the result of an induction of more rapid and greater overall metolachlor metabolism by the

ALS-inhibiting herbicides.

84

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