Pesticide Formulations - American Chemical Society

Chapter 5

Mode of Action of a Nonionic and a Cationic Surfactant in Relation to Glyphosate

H. de Ruiter, M. A. M. Verbeek, and A. J. M. Uffing

Downloaded by UNIV OF ARIZONA on October 11, 2016 | http://pubs.acs.org Publication Date: June 24, 1988 | doi: 10.1021/bk-1988-0371.ch005

Center for Agrobiological Research, P.O. Box 14, 6700 AA Wageningen, Netherlands

Addition of Ethomeen T/25 (polyoxyethylene (15) tallow amine) at d i f f e r e n t concentrations enhanced the glyphosate t o x i c i t y to seedlings of winter-wheat. The nonionic Renex 688 (polyoxyethylene (8) nonylphenol) showed a r e l a t i v e l y pore capacity to enhance glyphosate t o x i c i t y . Absorption of 14C-glyphosate is i n h i b i t e d by Renex 688 and enhanced by Ethomeen T/25, which explains the observed difference i n influence on glyphosate toxicity. Scanning electron microscopy revealed that the epicuticular waxes seem rather unimpaired a f t e r application of the surfactants. The two surfactants penetrate into the c u t i c l e . Application of Ethomeen T/25 resulted i n the appearance of necrotic lesions whereas Renex 688 gave much less injury. The r e s u l t s are discussed i n r e l a t i o n to the physical-chemical properties of Renex 688 and Ethomeen T/25.

Addition of r e l a t i v e l y high ( 0.1% (w/v)) concentrations of surfactant may give Increased phytotoxicity of glyphosate (8,12). The cationic polyoxyethylene amine surfactants appear to be the most appropriate compounds f o r combination with glyphosate (12). Because the influence of surfactants on glyphosate t o x i c i t y i s not completely understood (4,8), this study was started to develop a better understanding of the mode of action of surfactants. Insight i n the underlying mechanisms may lead to the use of surfactants based on r a t i o n a l arguments. 0

0097-6156/88/0371-0044$06.00/0 1988 American Chemical Society

Cross and Scher; Pesticide Formulations ACS Symposium Series; American Chemical Society: Washington, DC, 1988.


5.

DERUITERETAL.

Mode of Action ofa Nonionic and Cationic Surfactant

The experiments i n this study have been c a r r i e d out with the nonionic Renex 688 (polyoxyethylene (8) nonylphenol), the c a t i o n i c Ethomeen T/25 (polyoxyethylene (15) tallow amine) and technical grade glyphosate which have been applied to winter wheat grown under controlled environmental conditions. The difference between Ethomeen T/25 and Renex 688 i n their capacity to enhance glyphosate a c t i v i t y has been investigated by measuring the r e l a t i o n between surfactant concentration and glyphosate t o x i c i t y . As increased glyphosate t o x i c i t y may r e s u l t from an increased deposition, the r e l a t i o n between surfactant concentration and deposition was measured by fluorimetry. A f t e r passage of the c u t i c l e glyphosate i s thought to penetrate into the symplast (3). The c u t i c l e , the c e l l wall and the plasmalemma may be penetration l i m i t i n g factors for a water soluble anionic compound l i k e glyphosate as the c u t i c l e and the plasmalemma are l i p o p h i l i c and negatively charged. The c e l l wall i s permeable to water soluble compounds but also negatively charged. Addition of surfactants may r e s u l t i n increased absorption of glyphosate (4,7). It i s not obvious whether Increased permeability of the c u t i c l e or f a c i l a t e d entry into the symplast i s the main factor for t h i s phenomena. Scarce data on t h i s matter (8,11) give an i n d i c a t i o n that the f a c i l a t e d entry into the symplast may be the most relevant factor. In t h i s study the absorption of C-glyphosate i s measured at d i f f e r e n t concentrations of surfactant. Scanning electron microscopy has been used to investigate the capacity of the surfactants to dissolve e p i c u t i c u l a r waxes. Penetration of surfactants into the mesophyll must be preceded by d i f f u s i o n throughout the c u t i c l e . The absorption of Renex 688 and Ethomeen T/25 into the c u t i c l e has been measured by using a colorimetric method f o r determination. The influence of both surfactants on the permeability of the plasmalemma has been investigated by measuring the leakage of e l e c t r o l y t e s from potato discs a f t e r incubation i n surfactant containing medium. A rough estimation for penetration of surfactants into the tissue was obtained by observation of the appearance of necrotic l e s i o n s . Materials and Methods Plant Material. Winter wheat (cv. Arminda) was grown i n 11 cm-diam. p l a s t i c pots (6 plants/pot) f i l l e d with a mixture of sand and humic potting s o i l (1:2). The pots were subirrigated with ^-strength Steiner's nutrient solution (9). The plants were grown i n a growth chamber under the following conditions: 16 h l i g h t , 18/12 °C (day/night) temperature and 70% r e l a t i v e humidity. Light was provided by high pressure mercury lamps and incandescant lamps to give 65 W/m at l e a f l e v e l . 2

Phytotoxicity of glyphosate. The plants were treated at the 3-leaf stage. Sprays were applied with a laboratory sprayer f i t t e d with three nozzles (Birchmeier Hellco Sapphire 1.2 mm provided with a whirling pin 2F-0,6 mm perforated) to give a volume of 440 1/ha. Three weeks a f t e r treatment the parts above the ground were harvested by cutting at 1 cm above the ground and the fresh weight was measured.


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PESTICIDE FORMULATIONS: INNOVATIONS AND DEVELOPMENTS


Glyphosate a c t i v i t y i s expressed as percentage of the fresh weight of plants treated with demineralized water. Glyphosate solutions were prepared from technical grade glyphosate (isopropylamine s a l t ) . The application rate of the s a l t was equivalent to 0.095 kg glyphosate/ha. The surfactants were included i n the spray solution on a percentage (w/v) basis. Renex 688 (polyoxyethylene (8) nonylphenol) and Ethomeen T/25 (polyoxyethylene (15) tallow amine) were used i n t h i s study. Treatments were replicated four times i n a randomized complete block design i n a l l experiments• Deposition and surface tension measurements. Deposition of spray solutions (containing surfactants at appropriate concentrations) was quantified by spectrofluorimetry according to Richardson (6). Fifteen minutes a f t e r spray application the dye was washed o f f the plants. The absolute deposition i s expressed as the volume of spray solution retained per gram dry weight (ul/g). The r e l a t i v e deposition of the solutions i s expressed as percentage of the deposition of demineralized water. Treatments were replicated four times i n a randomized complete block design i n a l l experiments. The s t a t i c surface tension was measured according to the ring method. Three measurements were taken f o r each solution. 14 Absorption of C-glyphosate. After emergence the wheat seedlings were thinned to one seedling per pot. Applications were made at the 3-leaf stage. Methyl l a b e l l e d C-glyphosate (Amersham, s p e c i f i c a c t i v i t y 2.2 GBq/mmol) was converted to the monoisopropylamine s a l t by the addition of isopropylamine i n a 1:1 molecular r a t i o . Non-labelled technical grade glyphosate (monoisopropylamine s a l t ) , surfactants (on a weight to volume basis) and demineralized H^O were added to the C-glyphosate such that, the concentration of glyphosate (labelled plus non-labelled) amounts to 1.3 mM which i s the concentration used i n the spray solutions. The glyphosate solution was applied alone or i n combination with Renex 688 and Ethomeen T/25 as four 1-ul droplets (0.83 kBq) to a discrete area on the adaxial side of the second l e a f . The discrete area was marked by using waterproof drawing ink. A l l applications were made using a Burkard Microapplicator PAX 100 f i t t e d with a 50 u l syringe and PFTE coated needle. Each treatment was replicated three times i n a randomized complete block design. At indicated times the treated leaf was removed and washed with 5 ml water. This procedure removed 100% of the glyphosate immediately a f t e r droplet a p p l i c a t i o n . A 1-ml aliquot from the wash was added to 10 ml hydroluma (Lumac/3 M)• Radioactivity was quantified using standard l i q u i d s c i n t i l l a t i o n spectrometry techniques. Absorption of surfactants. Applications were made at the 3-leaf stage. Surfactant solutions (0.5% (w/v)) were applied as 10 1-ul droplets (50 ug) to a discrete area on the adaxial side of the second J.eaf using the microapplicator as described with the application of C-glyphosate. At indicated times the deposit was removed by gently wiping the treated area with a small p e l l e t of cotton-wool moistened with water. The p e l l e t was extracted with 6 ml chloroform. The amounts of surfactant were measured by using a colorimetric method (1)• Uniform plants were selected and each treatment was r e p l i c a t e d


5.

DE RUITERETAL.

Mode of Action ofa Nonionic and Cationic Surfactant

three times* The colorimetric method used showed a r e l a t i v e l y low s e n s i t i v i t y f o r Renex 688. Deposits smaller than 10 ug could not be measured accurately. Therefore the absorption of Renex 688 with deposits i n t h i s range i s expressed as "higher than 80%" (>80%).


Scanning electron microscopy (SEM). Conditions of application i n r e l a t i o n to the plants were as described f o r the absorption studies. Surfactant solutions at appropriate concentrations were applied as one 1-ul droplet. Twenty-four hours a f t e r a p p l i c a t i o n the leaf sections were removed from the plant and immedially frozen i n nitrogen slush and freezedried. Then the leaf sections were mounted on aluminium SEM stubs, sputtered with gold-palladium and examined i n the scanning electron microscope at an accelerating p o t e n t i a l of 15 kV. Leakage of e l e c t r o l y t e s . Potato discs (cv. Bintje) were cut (diam. 11 mm, thickness 2 mm) and washed twice with 20 ml water. Then 5 discs were transferred to a 100 ml f l a s k with 20 ml bathing medium containing surfactant solutions at appropriate concentrations. The flasks were gently shaken during one hour at 20 °C under natural illumination. Then the conductivity was measured and corrected f o r response caused by the bathing media themselves. The conductivity was expressed as percentage of the value obtained a f t e r incubation i n demineralized water. The incubations were r e p l i c a t e d three times i n one experiment. Estimation of i n j u r y . Conditions of application i n r e l a t i o n to the plants were the same as described f o r the absorption studies. Surfactant solutions at appropriate concentrations were applied as three 1-ul droplets. Seventy-two hours a f t e r application the i n j u r y was estimated by v i s u a l observation of the appearance of necrotic lesions. Results and Discussion Glyphosate phytotoxicity. The low application rate of glyphosate gave a small reduction i n fresh weight (Figure 1). Ethomeen T/25 and Renex 688 enhanced the glyphosate t o x i c i t y . At a l l concentrations applied Ethomeen T/25 gave a higher reduction than Renex 688. Addition of Ethomeen T/25 gave a rapid reduction i n fresh weight. With Renex 688 the phytotoxicity does not change when the surfactant concentration increased from 0.001% (w/v) to 0.01% (w/v). The r e s u l t s with Ethomeen T/25 agree with the observation that addition of MON 0818 (polyoxyethylene tallow amine) at d i f f e r e n t concentrations enhanced the glyphosate t o x i c x i t y to seedlings of common milkweed and hemp dogbane (12). The r e l a t i v e l y pore capacity of Renex 688 to enhance glyphosate phytotoxicity agrees to a c e r t a i n extent with the observation that a s i m i l a r surfactant (formulated as Agral 90) does not influence the glyphosate t o x i c i t y to several species (5). A p p l i cation of the surfactants themselves at 1.0% (w/v) d i d not result i n reduction of fresh weight.

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Cross and Scher; Pesticide Formulations Washington, D.C. 20036 ACS Symposium Series; American Chemical Society: Washington, DC, 1988.

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Deposition. With the discussion on the deposition data the assumptions have been made that the behaviour of the fluorescent dye i s the same as that of glyphosate and that the presence of glyphosate (1.3 mM) does not change the data obtained with surfactant solutions. Both surfactants increase the deposition on the d i f f i c u l t to wet wheat seedlings (Figure 2). At a surfactant concentration of 1% (w/v) a 5 to 6 times enhanced deposition could be observed. The r e l a t i o n between concentration and deposition i s similar f o r Ethomeen T/25 and Renex 688 with Renex 688 giving a somewhat higher deposition. As a difference i n deposition does not seem to cause the observed difference i n phytotoxicity (Figure 1), difference i n entry of glyphosate into the plant must give the difference i n phytotoxicity. By taking the point at which the surface tension did not further decrease (data not shown) the c r i t i c a l micelle concentrations (CMC) of the surfactants can be estimated. The CMC amounts to 0.003% (w/v) for Renex 688 and 0.01% (w/v) f o r Ethomeen T/25. This implies that the continuously increasing deposition at r e l a t i v e l y high concentra tions of surfactant can not be explained by decreased s t a t i c surface tension. I t was suggested by Taylor (10) that r e l a t i v e l y high concen trations of surfactant are necessary as the concentration of surfac tant a f f e c t s the speed at which the surface tension of the drop i s changed during the time the drop moves from the nozzle towards the plant• 14 Absorption of C-glyphosate. The absorption data of glyphosate alone and glyphosate with Ethomeen T/25 at 0.05% (w/v) and 0.5% (w/v) show that most r a d i o a c t i v i t y has been absorbed within 6 h after a p p l i c a t i o n (Table I ) . a 14 Table I. Absorption of C-Glyphosate % of applied Dose Treatment Glyphosate Glyphosate + Glyphosate + Glyphosate + T/25 0.05% Glyphosate + T/25 0.5 %

6h

24h 20.6

48h

Renex 688 0.05% Renex 688 0.5 % Ethomeen

21.5 2.5 11.8 49.2

± 3.1 ±0.9 ± 2.4 + 1.8

± 4.2 17.8 ± 1.5 54.1 ± 5.0

31.6 ± 7.4 9.0 ± 1.9 23.1 ± 2.6 67.6 ± 3.3

Ethomeen

46.5

± 3.6

60.1

61.7

± 2.2

±

3.8

^Means ± SE calculated from 2 experiments with each 3 r e p l i c a t e s - • no absorption Ethomeen T/25 at 0.05% (w/v) and 0.5% (w/v) enhances the absorption whereas Renex 688 at 0.05% (w/v) and 0.5% (w/v) gives a decreased absorption. This observation confirms the conclusion derived from the phytotoxicity and the deposition data with respect to a possible l i m i t e d entry of glyphosate i n the presence of Renex 688. A more than twofold Increased glyphosate absorption as the result of addition of a polyoxyethylene tallow amine (MON 0818) was also observed with f i e l d bindweed (7). An i n h i b i t o r y e f f e c t on glyphosate absorption was


5.

DERUITERETAL.

Mode ofAction of a Nonionic and Cationic Surfactant


• « Ethomeen T/25 o* Renex 688

20 10 0 10"

1.0 10" 10ri surfoctont concentration (%w/v)

Figure 1. Effect of surfactant concentration on glyphosate t o x i c i t y expressed as % of the control's fresh weight. Values are means ± SE calculated from 4 experiments with each 4 r e p l i c a t e s . The control's fresh weight (mean value per experiment) ranged from 32 to 46 g per pot.

700

• « Ethomeen T/25 o*Renex 688

600 -

a 200 100

0.01 0.05 0.1 05 1.0 surfactant concentration (% w/v)

Figure 2. Effect of surfactant concentration on deposition. Deposition i s expressed as Z of the control sprayed with water. Values are means ± SE calculated from 3 experiments with each 4 r e p l i c a t e s . Deposition of the control (mean value per experiment) ranged from 41 to 54 u l / g dry weight.


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demonstrated with Renex 36 (polyoxyethylene 6- t r i d e c y l ether) after application to barley seedlings (4). Based on enhanced leakage of e l e c t r o l y t e s from barley segments the suggestion was made that disruption of membrane i n t e g r i t y may lead to reduced absorption (4). Our observation that lowering of the concentration of Renex 688 from 0.5% (w/v) to 0.05% (w/v) results i n a higher i n h i b i t i o n of glyphosate absorption seems to c o n f l i c t with t h i s suggestion. Table I I . Absorption

of Surfactants


% of applied Dose 6h

24h

0.5% (w/v)

62.8 ± 7.8

> 80

Ethomeen T/25 0.5% (w/v)

38.4 ± 7.2

56.0 ± 5.4

Renex 688

48h

b

> 80

b

73.2 ± 3.5

^Means ± SE calculated from 3 experiments with each 3 r e p l i c a t e s See section Materials and Methods Absorption of surfactants. Renex 688 and Ethomeen T/25 penetrate into the c u t i c l e (Table I I ) . With both surfactants more than a h a l f of the deposit i s absorbed within 6 hours a f t e r a p p l i c a t i o n . The course of uptake and the percentages of uptake agree along rough l i n e s with the r e s u l t s obtained a f t e r application of C-polyoxyethylene tallow amine (MON 0818) to f i e l d bindweed (8) and polyoxyethylene (8.5) nonylphenol to wheat (2). The values f o r absorption of Renex 688 exceed those f o r Ethomeen T/25. Hydration of both surfactants at 95% RH (data not shown) revealed that hydrated Renex 688 e x i s t s as a g e l whereas hydrated Ethomeen T/25 shows a soft s o l i d form. These differences i n uptake and f y s i c a l form support the conclusion of Anderson and G i r l i n g (2) that the uptake of surfactants i s at least p a r t l y related to t h e i r physical form. The rapid penetra t i o n of Renex 688 into the c u t i c l e does not provide an obvious explanation f o r the observed i n h i b i t i o n of glyphosate absorption. The movement of Ethomeen T/25 into the c u t i c l e implies that increased c u t i c l e permeability, injury to underlying c e l l s and/or complexation with glyphosate may lead to the observed increase i n absorption (Table I ) . Scanning electron microscopy. Scanning electron micrographs (Figure 3) show that the c r l s t a l l i n e e p i c u t i c u l a r waxes seem rather unimpaired 24 h a f t e r a p p l i c a t i o n of surfactants. After a p p l i c a t i o n of Ethomeen T/25 at 0.05% (w/v) and 0.5% (w/v) a layer of surfactant covering the e p i c u t i c u l a r waxes can be observed. F i t s are v i s i b l e (Figure 3F) which may be the result of small a i r pockets within the deposit which burst during drying. Application of Renex 688 at 0.05% (w/v) and 0.5% (w/v) r e s u l t s i n a much thinner deposit. This results confirms the rapid penetration of Renex 688 (Table I I ) . The results with Ethomeen T/25 concur with scanning electron micrographs obtained a f t e r a p p l i c a t i o n of polyoxyethylene tallow amine (MON 0818) to f i e l d bindweed (8)• A deposit of surfactant was observed and there was no


5.

DERUITERETAL.

Mode ofAction ofa Nonionic and Cationic Surfactant


evidence f o r disruption of e p i c u t i c u l a r waxes. These r e s u l t s do not form a proof but indicate that enhanced absorption of glyphosate i n the presence of Ethomeen T/25 i s not a r e s u l t from dissolved c u t i c u l a r waxes. With Ethomeen T/25 at 0.5% (Figure 3E) and to a smaller extent at 0.05% (w/v) a shrinking of underlying epidermal c e l l s could be observed which indicates disruption of the plasmalemma by Ethomeen T/25 within 24 h a f t e r a p p l i c a t i o n . Application of Renex 688 d i d not result i n shrinking of c e l l s . This observation and the observed penetration of Renex 688 into the c u t i c l e (Table II) lead to the conclusion that Renex 688 does not diffuse away from the underside of the c u t i c l e or Renex 688 i s less disruptive than Ethomeen T/25 to the plasmalemma. Leakage of e l e c t r o l y t e s . Measurement of e l e c t r o l y t e leakage from leaf discs pretreated with surfactant or a f t e r incubation i n surfactant containing medium may be affected by the c u t i c l e as a b a r r i e r f o r penetration of surfactants into the tissue. For that reason potato discs were used i n t h i s study. Ethomeen T/25 and Renex 688 enhanced the leakage of e l e c t r o l y t e s (Table I I I ) . A r e l a t i v e l y large increase of the leakage can be observed at a r e l a t i v e l y low concentration of surfactant (0.005% (w/v)). The leakage with Ethomeen T/25 exceeds the leakage with Renex 688 at a l l concentrations but the values are of the same of order of magnitude. Table I I I . Leakage

of E l e c t r o l y t e s from Potato Discs Conductivity as % of Control

Surfactant

Concentration of Surfactant 0.05% (w/v) 0.005% (w/v) 0.05% (w/v)

Renex 688

343

454

517

Ethomeen T/25

506

846

1050

*Mean values from a representative experiment with 3 r e p l i c a t e s The membrane disrupting property of Renex 688 and the absence of shrinked epidermal c e l l s as was observed with scanning electron microsopy support the idea that Renex 688 penetrates r a p i d l y into the c u t i c l e but does not or only to a small extent diffuse from the c u t i c l e to the apoplast. Injury caused by surfactants. Application of Ethomeen T/25 at 0.05% (w/v) and 0.5% (w/v) gave necrotic lesions (observed 72 hours a f t e r a p p l i c a t i o n ) . At 0.5% (w/v) t h i s injury was more severe than at 0.05% (w/v). Renex 688 d i d not give injury symptoms at 0.05% (w/v) but at 0.5% (w/v) some small necrotic lesions could be observed. This result supports the view that Renex 688 i s retained by the c u t i c l e . Never theless at the r e l a t i v e l y high concentration of 0.5% (w/v) penetra tion of Renex 688 occurs which may p a r t l y explain the enhanced glyphosate absorption compared with absorption at 0.05% (w/v) (Table I).


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Figure 3. Scanning electron micrographs of the adaxial side of winter wheat; A) Untreatted, x 10 000; B) Renex 688 0.05% (w/v), x 10 000; C) Renex 688 0.5% (w/v), x 10 000.


DERUITERETAL.



5.

Figure 3. Continued D) Ethomeen T/25 0.05% (w/v), x 10 000; E) Ethomeen T/25 0.5% (w/v), x 200; F) Ethomeen T/25 0.5% (w/v), x 10 000.


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PESTICIDE FORMULATIONS: INNOVATIONS AND DEVELOPMENTS Table IV. Summarized Data from this and two other Studies

Surfactant

Ethomeen T / 2 5 Renex 688 Tween 20, Renex 36 Tween 2 0 ° MON 0818

a

a


C

a

HLB

Ionig form

19.3 12.3 16.7 11.5 16.7

C N N N N C

t h i s study; O'Donovan JT a.o. C» c a t i o n i c , N « nonionic.

Species

wheat wheat barley barley field bindweed

Enhanced Glyphosate absorption

(4,7) Injury

+

+

+

+

-

+

+

(1985); ° S h e r r i c k SL a.o.

+ (1986);

Conclusions The absorption of glyphosate i s i n h i b i t e d by Renex 688 and enhanced by Ethomeen T/25. Both surfactants are absorbed by the c u t i c l e . The r e s u l t s of t h i s study Indicate that the p a r t i t i o n of these surfactants between the l i p o p h i l i c c u t i c l e and the apoplast favours a p o s i t i o n of Renex 688 i n the c u t i c l e whereas Ethomeen T/25 diffuses to a larger extent than Renex 688 into the apoplast. This suggestion agrees with the l i p o p h i l i c character of Renex 688 and the high water s o l u b i l i t y of the cationic Ethomeen T/25. The presence of Renex 688 i n the c u t i c l e may cause the observed i n h i b i t i o n of glyphosate absorption when Renex 688 i s appllcated at 0.05% (w/v). However the cause of the p a r t i a l n e u t r a l i z a t i o n of this i n h i b i t i o n with Renex 688 at 0.5% (w/v) i s not c l e a r . The penetration of Ethomeen T/25 into the underlying tissue may lead to Increased permeability of the c e l l wall and the plasmalemma to glyphosate. The p o s s i b i l i t y of c o r r e l l a t i o n between enhanced absorption of glyphosate and penetration of surfactants into the tissue i s also supported by two other studies (4,7) on this matter (Table IV). Acknowledgments The authors acknowledge AKZO Chemie for providing Ethomeen T/25, ICI-Holland for providing Renex 688 and B . V . Luxan for providing technical grade glyphosate. Literature Cited 1. 2. 3. 4. 5. 6.

Anderson, N . H . ; G i r l i n g , J. Analyst 1982, 836-838. Anderson, N . H . ; G i r l i n g , J. P e s t i c . S c i . 1983, 14, 399-404. Gougler, J.A.; Geiger, D.R. Plant P h y s i o l . 1981, 68, 668-672. O'Donovan, J.T.; O ' S u l l i v a n , P . A . ; Caldwell, C.D. Weed Res. 1985, 25, 81-86. O ' S u l l i v a n , P . A . ; O'Donovan, J.T.; Hamman, W.M. Can. J. Plant S c i . 1981. 61, 391-400. Richardson, R . G . Australian Weeds 1984, 3, 123-124.


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DERUITERETAL.


7. Sherrick, S.L.; Holt, H.A.; Hess, F.D. Weed S c i . 1986, 34, 811-816. 8. Sherrick, S.L.; Holt, H.A.; Hess, F.D. Weed S c i . 1986, 34, 817-823. 9. Steiner, A.A. ISOSC Proc. 6th Int. Congress on S o i l l e s culture 1984, 633-650. 10. Taylor, W.A.; Shaw, G.B. P e s t i c . S c i . 1983, 14, 659-665. 11. W y r i l l , J.B.; Burnside, O.C. Weed S c i . 1976, 24, 517-566. 12. W y r i l l , J.B.; Burnside, O.C. Weed S c i . 1977, 25, 275-287. December 28, 1987


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Pesticide Formulations - American Chemical Society

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