Pestic. Sci.

1993, 38, 227-235

The Role of Surface Tension of Spreading

Drodets in Absorption of a Herbicide
Formulation via Leaf Stomata*
Rosalind D. Buick,"$ Graeme D. Buchanb & Roger J. Fielda$
"Plant Science and *Soil Science Departments, Lincoln University, Canterbury, New Zealand
(Revised manuscript received 31 July 1992; accepted 21 May 1993)

Abstract: The foliar absorption by field bean (Viciu fubu L.) of triclopyr-
triethylamine from solutions containing an organosiiicone surfactant, ' Silwet L-
77' at O, 1.0, 2.5 and 5.0ml litre-l, was investigated using fluorescence and
radiotracer studies. Physical characteristics such as formulation surface tension
and contact angles of droplet formed on the field bean leaf were determined and
combined to obtain a critical surface tension value (yCrltlpa,)of 19.5-22.9 mN m-'
for the adaxial leaf surface. The apparent surface tensions of the three formulations
containing L-77 were equivalent and low, an average of 22 mN m-l, which would
be sufficiently low to permit stomatal infiltration. The presence of L-77 at 2.5 or
5.0 ml litre-' significantly enhanced the uptake of triclopyr-triethylamine into
the leaf, particularly via stomatal pore regions. However, stomatal uptake from
solutions containing 0 or 1.0 ml litre-' L-77 was limited or non-existent. A
phenomenon termed 'adsorptive dilution ' is proposed to explain the absence of
stomatal infiltration with the 1.0 ml litre-' formulation. This involves a migration
of surfactant from the bulk solution to the expanding droplet surfaces as the
droplet spreads. This dilutes the surfactant concentration within the bulk solution,
which can in turn increase surface tension.

1 INTRODUCTION solution combination. Schonherr and Bukovacl found a

The entry of spray solutions into a plant can occur via
two main physical pathways: through the cuticle or
through stornatal pores. Conditions critical for achieving
stornatal infiltration of solutions were identified by
Schonherr and Bukovac.' The stomatal pore wall
morphology must be such that the contact angles formed
between the solution and the pore walls are conducive to
a positive pressure difference inducing liquid movement
into the substomatal cavity. The solution surface tension
is also important and needs to be less than the critical
surface tension value (ycritical)for that plant surface and
* Paper based in part on material presented at the 'Third
International Symposium on Adjuvants for Agrochemicals',
organised by the Physicochemical and Biophysical Panel of the
SCI Pesticides Group and held at the University of Cambridge,
Cambridge, UK on 3-7 August 1992.
t Present address : c/o Department of Entomology, Virginia
Polytechnic Institute & State University, Blacksburg, Virginia
24061-0319, USA.
0 To whom correspondence should be addressed.
227
Pestic. Sci. 0031-613X/93/$06.00 0 1993 SCI. Printed in Great Britain
critical surface tension value of 30 m N rn-l for Zebrina
purpusii Bruckn, only some surfactants being able t o
reduce suface tension below this value. Greene and
Bukovac2observed stornatal penetration into leaves only
when surface tension was lowered sufficiently by certain
surfactants in the formulation.
Surfactants are made u p of surface-active molecules,
with a hydrophilic and a hydrophobic p ~ r t i o n .The ~
ability of a surfactant to reduce surface tension depends
on the hydrophilic-hydrophobic relationships and the
propensity of the molecules to arrange themselves at the
interfaces of different phases (i.e. between the droplet
and the air and/or between the droplet and the plant
surface).4
The critical micelle concentration (CMC) of a sur-
factant in solution occurs a t the break point or inflexion
point in the plot of surface tension against the logarithm
of c ~ n c e n t r a t i o n .At
~ and above this point, surfactant
monomers in solution will typically aggregate into
colloid-type structures known as m i c e l l e ~ . ~Micelle
228
formation has been known to cause marked changes in
surfactant characteristia6 However, this may be of less
importance with organosilicone surfactants such as
' Silwet ' L-77 since their micelle formation is very
different to that of the traditional surfactants.?
L-77 is an organosilicone surfactant known for its
dramatic effect in reducing surface tension of spray
solutionss, and its ability to enhance the performance of
herbicides in the control of certain weed species."-" The
mode of action of L-77 has been linked to the promotion
of herbicide uptake via stomatal pores.'O' "-15
Many researchers have observed the lack of correlation
between surface tension and wetting characteristics of
herbicide formulations and their perf~rmance.~, 16, l 7 In
this study the surface tension and wetting characteristics
of a triclopyr-triethylamine formulation with various
concentrations of the organosilicone surfactant, L-77,
were investigated and related to absorption by field bean
(Vicia fuba L.) leaves. This species was selected as a
model plant species for gorse (Ulex europaeus L.), a
significant woody weed problem in New Zealand. Field
bean's uptake behaviour with a number of triclopyr-
organosilicone formulations was found to be similar to
that of gorse, and a number of similarities in stomatal
architecture between gorse and field bean were observed.
Field bean grows rapidly, and is very suitable for uptake,
translocation and many physicochemical studies.ls
The use of a Zisman plot enabled the critical surface
tension value for the field bean leaf surface to be
determined and hence those triclopyr formulations likely
to infiltrate stomatal pores could be predicted. An
innovative interpretation of the results for surfactant
behaviour within droplets enabled some interesting
conclusions to be drawn with regard to herbicide
performance, surface tension, and L-77 concentration.
In particular, calculations reveal the importance of an
'adsorptive dilution' effect, whereby the increase in
droplet surface accompanying spreading causes sur-
factant migration from the bulk to the surface, thus
lowering the effective surfactant concentration within the
bulk film. This in turn can alter the surface tension and
hence affect stomatal penetration.
2 MATERIALS AND METHODS
2.1 Plant material
The field bean (Vicia faba L. cv. Maris Bead) seeds were
pregerminated at 25°C on moist blotting paper for three
to four days prior to sowing. Plants were grown in 120-
mm pots containing potting mix for two weeks in a
controlled environment cabinet (day/night temperatures
21/15"C; 14: 10 h1ight:dark; 70-75% relative humidity;
light intensity 300 pmoles m-2 s-l). All formulation
applications to field bean plants were made to adaxial leaf
surfaces of the third leaf once it had fully expanded.
R. D. Buick, G . D. Buchan, R. J . Field
2.2 Fluorescence microscopy
Solutions of triclopyr-triethylamine with L-77 at 0, 1.0,
2.5, and 5.0 ml litre-' plus 'Uvitex' 2B (Ciba-Geigy), a
fluorescent indicator, at 1 g litre-' were prepared. Leaves
were treated with 4 ~ 0 . pl 5 droplets, 5-10 mm away
from the leaf midrib. At selected times (1 and 60 min),
treated leaves were first rinsed with deionised water, then
with methanol+water (70+30 by volume). The two
rinses were carried out to ensure that no herbicide
residue remained on the leaf surface. Rinsed leaves were
gently dried with clean tissue and mounted onto glass
slides for observation. Herbicide solutions were detected
using a Zeiss fluorescence microscope with a UV
excitation source and representative photomicrographs
were obtained of herbicide absorption within the droplet
area.
Image analysis was then used to quantify the mean
areas of herbicide absorption in each photomicrograph
as shown by the fluorescent indicator. The system was an
image analysis package (PC-Semper, Synoptics Ltd.,
Cambridge, UK) with attached camera and video
monitors, capable of scanning images of up to 512 x 512
pixels with 256 grey levels. Threshold intensities were
selected to obtain a scanned video image as close to the
micrograph as possible. The mean values for degree of
herbicide absorption were expressed for each treatment
as (1) area of absorption as a percentage of whole
photomicrograph area, and (2) absorption area (mm2)in
whole photomicrograph area.
Transformations were performed on the data to
validate the analysis of variance assumption of equal
variances. A square root transformation of the area
(mm') and a logit transformation ( y = loglo(x/(100-x)))
on the percentage area were the most
suitable. As statistical inferences were made on the
transformed data, standard errors of the transformed
means are given (Table 1).
2.3 Foliar uptake studies
['4C]Triclopyr-triethylamine solutions were made by
adding [I4C]triclopyr to a solution of triclopyr-
triethylamine (360 g a.e. litre-') made up at a rate
equivalent to 4 kg a.e. ha-' in 200 litre ha-'. Relative
concentrations of labelled and unlabelled triclopyr
were such that 1 pl of herbicide solution contained
20000-30000 dpm.
Four formulations of amine were prepared with
surfactant concentrations of 0, 1.0, 2.5, and 5.0 g litre-'.
The single microlitre of formulation for each plant was
applied as 4 x 0 . 2 5 ~ droplets,
1 2 h after the beginning of
the light photoperiod. Eight replicate plants per treatment
were used. After 1 or 6 h, bean leaves were washed with
+
methanol + water (70 30 by volume) to determine the
percentage gross uptake. Immediately after the methanol
wash, a second leaf wash with 10 ml of chloroform was
Surface tension in absorption of herbicide via leaf stomata 229

TABLE 1 2.4 Surface tension

Image Analysis Results from Fluorescence Photomicrographs
showing Means for Individual Treatments Solutions of triclopyr-triethylamine were made up in
deionised water at a rate equivalent to 4 kg a.e. ha-' in
L-77 Time Mean absorption" Mean absorption 200 litre ha-'. Various concentrations of L-77 were
(ml litre-l) (min) (YOof total area) areaa added to these amine solutions; 0, 0.05, 0.1, 0.5, 1.0, 2.5
(mm2) and 5.0 ml litre-l. All measurements were carried out at
0 1 094 0.19 22°C.
( - 2.02, 0.52) (0.43, 0.75) Surface tension was measured under equilibrium
60 1.oo 0.20 conditions using an adapted droplet-volume technique,
( - 2.07, 0.30) (0.43, 0.43) as described by Taylor.''. l9 Twenty replicate droplets
1.o 1 0.52 0.10 were formed from the apparatus for each solution.
(-2.30, 0.36) (0.56, 0.37) Results were plotted against both linear and logarithmic
60 1.57 0.3 1 scales of surfactant concentration. Results for L-77
( - 1.81, 0.26) (0.56, 0.37) solutions in water were also plotted to compare with the
2.5 1 8.26 1.65 triclopyr-triethylamine formulations. Linear regression
(- 1.18, 0.19) (1.19, 0.28)
analysis was used to describe the data in the surface
60 39.28 7.78
(-0.43, 0.19) (2.50, 0-28) tension-log concentration plots.
5.0 1 12.65 2.53 The effect of including the fluorescent indicator was
(-0'86, 0.23) (1.57, 0.33) determined by measuring surface tension of four
60 33.62 6-73 triclopyr-triethylamine and L-77 solutions (0, 1.0, 2.5,
(-0'32, 0.30) (2.55,0.43) and 5.0 ml litre-l L-77) with and without 'Uvitex' 2B at
1 g litre-'.
a Values in parentheses are transformed mean percentage
absorption (logit) or transformed mean absorption area (square
root), followed by standard errors of the transformed means. 2.5 Contact angles (0,) and zisman plot

Advancing contact angles (0,) of herbicide droplets on

used to determine the amount of radiolabel in the wax as
a percentage of total applied radioactivity. Percentage
net uptake of triclopyr-triethylamine was determined by
subtracting the wax uptake value from gross uptake.
Droplets (0.25 pl) of triclopyr-triethylamine applied to
field bean leaves appeared to be dry after 6&120 s if no
surfactant was present, and within 60 s if L-77 was
present in the formulation."
Arcsine transformations on the percentage uptake
data were found to best equalise the standard errors. All
statistical inferences were based on the transformed
means and standard errors of the means, but non-
transformed means and errors are also given (Table 2).
adaxial surfaces of bean leaves were recorded using a
horizontally oriented travelling microscope with an
attached micrometer eyepiece. The solution concentra-
tions used were identical to those used in the surface
tension measurements. Ten replicate droplets per sol-
ution were formed. Temperature and relative humidity
were maintained at 22°C and 7&75% respectively.
Measurements were always recorded 10 s after drops
were applied to the leaf.
Cosines of the contact angles were plotted against
surface tension values for each solution to obtain a
Zisman plot.20The critical surface tension (ycritica,)for
the field bean adaxial leaf surface could then be

TABLE 2
Percentage Uptake (Gross, Wax and Net) of [14C]triclopyr-triethylamine
after 1 h and 6 h when Concentration of L-77 in the
Solution was Increased"

L-77 I hour 6 hours

(ml litre-')
Gross (%) Wax (Yo) Net (YO) Gross (YO) W a x (YO) Net (%)

0.8 (4.9) 0.2 (2.6) 0.6 (4.2) 11.5 (19.6) 4.9 ( 12.4) 5.0 ( 12.6)
2.8 (9.5) 0.3 (3.0) 2.6 (9.0) 12.4 (20.4) 2.5 (8.7) 10.2 (18.5)
6.8 (14.7) 0.3 (3.3) 6.5 (14.3) 17.1 (24.4) 1.6 (7.0) 15.9 (23.5)
9.7 (18.1) 0.3 (3.2) 8.8 (17.2) 16.4 (23.8) 1.4 (6.7) 15.0 (22.8)
l.OO(l.11) 0.03 (0.14) 0.99 (1.15) 1.31 (1.12) 0.54 (0.92) 1.10 (1.02)

Arcsine transformed means are given in parentheses and standard errors (SE,) below.
230
determined by extrapolating a line of best fit for the
plotted data, back to where cosine 6, = 1 (see Fig. 3).
3 RESULTS
3. I Fluorescence microscopy
After both 1 and 60min washes, the control herbicide
solutions (i.e. no added L-77) had not penetrated the
adaxial surface of field bean leaves, shown by a relative
absence of blue fluorescent dye across their surfaces. The
glandular hairs or trichomes and any dust particles on
the leaves were the only parts which appeared to absorb
the fluorescent indicator and presumably the herbicide.
When L-77 was added to the triclopyr-triethylamine
solution at 1.0 ml litre-' there were some small and
infrequent regions of herbicide absorption, mainly
associated with trichomes (after 1 and 60 min), but in
some cases with stornatal pores after 60 min. By contrast,
L-77 at 2.5 or 5.0 ml litre-' both induced similar extensive
regions of herbicide uptake mainly associated with
stornatal pores, both after 1 and 60 min. Herbicide
damage to the leaf epidermal cells after 60 min with the
two highest L-77 concentrations was characterised by
large and brightly lit regions of indicator completely
covering the site of droplet application.
Image analysis found the mean area of triclopyr
absorption was greater after 60 than after 1 min with all
treatments (Table 1). Average absorptions as a per-
centage of total photograph area for 1 and 60min
treatments were 6.0 and 18.9% respectively. The dif-
ference between the times was most notable for 2.5 and
5.0 ml litre-' L-77 treatments (Table 1).
Herbicide absorption areas were significantly greater
when 2 5 or 5.0 ml litre-' L-77 was added to the triclopyr-
triethylamine solution. These two treatments were not
significantly different from each other (Table 1). Similarly
the control and 1.0 ml litre-' L-77 treatments were not
.,i
-E 50
2ok ' 1' ' 2* ' 3' ' 4*
' 5
I
'Silwet' L77 concentration (ml litre.')
Fig. 1. Surface tension of triclopyr-triethylamine solutions
containing 'Silwet' L-77. Standard error of the mean = 0.71.
R . D . Buick, G . D . Buchan, R . J . Field
significantly different from each other. The interaction
between time and solution treatments was not significant.
A range of herbicide absorption levels was often
detected for a single treatment. This was reflected in the
large standard errors for some treatments in Table 1.
3.2 Foliar uptake studies
Gross and net uptake of ['4C]triclopyr-triethylamine was
significantly increased above control after 1 h by the
inclusion of L-77 at any of the three concentrations
(Table 2). Incorporation into wax increased as the rate of
L-77 increased and was only significantly greater than
control when the two higher concentrations were used
(i.e. 2.5 or 5.0 ml litre-') (Table 2).
After 6 h, gross and net uptake of triclopyr-
triethylamine was also increased by the addition of L-77
at all three concentrations. However, this increase was
only significantly above control with the 2 5 ml litre-'
concentration for gross uptake and with any of the three
concentrations for net uptake. Of these three L-77
treatments, the 2.5 and 5.0 ml litre-' concentrations
significantly improved net uptake above the value
with 1.0 ml litre-' surfactant. Incorporation into wax
decreased as the rate of L-77 increased, such that at the
two highest surfactant concentrations, incorporation
into the wax was significantly less than that found in the
control. These results are discussed below in terms of
selecting an optimum concentration of L-77 to maximise
the foliar uptake of triclopyr-triethylamine.
3.3 Surface tension
The addition of a fluorescent indicator, 'Uvitex' 2B, did
not significantly alter the surface tension of any of
the triclopyr-triethylamine + L-77 solutions (data not
shown). The surface tension of the triclopyr-
triethylamine solution with no added surfactant was 63.4
(k1.4) mN m-' but this was significantly reduced by the
addition of 'Silwet' L-77 (Fig. 1). At concentrations
greater than 0.5 ml litre-' there were no significant
changes in surface tension from a mean of 22.8 mN m-'.
Thus, the largest decrease in surface tension occurred
with the addition of L-77 up to 0.5 ml litre-'. Surface
tension values were also plotted against logarithm of
surfactant concentration for L-77 in aqueous solution
and in the triclopyr-triethylamine solution (Fig. 2). A
double line or 'broken-stick' trend was obtained in both
cases. The right-hand observations were regressed, while
the left-hand mean observations (0.05 and 0 1 ml litre-')
were simply joined. In the case of L-77 in water solutions,
the regression equation was :
y = 22.6 - 3 . 0 9 ~ (R2= 62 Yo)
The triclopyr-triethylamine and L-77 formulations
formed a regression line described by :
y = 22.5-0482~ (R2= 31 O h )
Surface tension in absorption of herbicide via leaf stomata
50r
451
r for the adaxial leaf surface of field bean was calculated as
I
10' 100 10'
'SiIwet' ~ 7 concentration
7 (ml litre-')
Fig. 2. Surface tension of solutions of 'Silwet' L-77 in (A)
water and in ( 0 )triclopyr-triethylamine solutions (log scale).
Bars = least significant differences. Linear regression to the
surface tension results for 0.5, 1.0, 2.5 and 5 0 m l litre-'
L-77 solutions in water was: y = 2 2 6 3 . 0 9 ~(R2= 62%);
in triclopyr-triethylamine solution was : y = 22.5-0.482~
(R2= 31 %).
Surface tension (mN m-')
Fig. 3. Relationship between cosine of the contact angles of
droplets formed on the adaxial leaf surfaces of field bean and
surface tension (mN m-l) of the triclopyr-triethylamine L-77 +
solutions. The mean critical surface tension value (yCritica,)value
was 21.1 mN m-l, with 95% confidence limits shown as the
shaded region.
The intercept of the two lines was at approximately
0.2 ml litre-' L-77 in both cases. The limiting slope (i.e.
slope of the steeper line) and the intercept of the two lines
are discussed in terms of surface adsorption, area per
molecule, droplet thickness and critical micelle con-
centration (CMC) calculations.
3.4 Contact angles
A quadratic curve gave the best fit to the relationship
between surface tension and cosine of contact angle:
R2 = 83 YO(Fig. 3). The equation was:
y = 2.09 - 0.0655~ + 0.000667~~
23 1
where x = surface tension in mN m-l and y = cosine of
contact angle (8,).
From this, the critical surface tension value (ycritica,)
21-1 mN m-l. Parallel curves fitted to provide 95%
confidence limits yielded a ycritical range of
19.5-22.9 mN m-' (Fig. 3). The implications and suit-
ability of the quadratic model are discussed below.
3.5 Adsorptive dilution theory
This section addresses the problem posed in the above
results by the unexpectedly low uptake of herbicide from
solutions containing 1.0 ml litre-' L-77, despite the fact
that the corresponding surface tension should have been
below the critical value for stomata1 entry.
Surfactant molecules tend to aggregate at the interface
of different phase^.^.'^^^ Depending on the propensity of
surfactant molecules to arrange themselves at these
interfaces (which here include the solution-air and
solution-leaf interfaces), any increase in interface area
will produce a reduction in surfactant Concentration
within the bulk solution. The greater the spread of the
droplet, the greater will be the reduction in bulk
concentration. Here, the known details of the surface
adsorption of L-77 from solution were used in first-order
calculations (1) to establish that the film produced from
droplet spread was sufficiently thick to be considered
'macroscopic' within its bulk, and (2) to determine the
effective change in surfactant concentration within the
bulk solution (specifically for a droplet of 1.0 ml litre-'
L-77 close to the critical micelle concentration).
3.5.1 Macroscopic nature of theJilm
First it is necessary to establish that the droplet film
formed on the field bean leaf was of 'macroscopic'
thickness. If the film is macroscopic (i.e. with thickness
much greater than the average distance between sur-
factant molecules in solution), then the surfactant
solution can be considered to behave like 'bulk' solution,
so macroscopic concepts of a bulk concentration paired
to its corresponding surface tension can be applied.
Mean droplet thickness (1) was calculated from eqn (1) :
t = V/A (1)
= 2.5 x m3/37 x m2
= 6.8 x m
where V = volume of droplet applied (0.25 p l )
and A = apparent area of droplet spread, found to
be 37.0 mm2.1591s
For present purposes, the average intermolecular
distance between surfactant molecules was calculated for
a concentration of 0.2 ml litre-', close to the apparent
232
critical micelle concentration for L-77 in water.' If the
macroscopic assumption holds at this low concentration,
then it would apply even more so at higher concentra-
tions, such as 0.5 or 1.0 ml litre-', for which the droplet
spreads were found to be e q ~ a 1 . l ~
Consider a total volume, K, of a solution of con-
centration, C. The volume of surfactant, E,in the volume
of water, V,, determines the concentration.
i.e. I/I= K+V,
and c =y / y
The mass of L-77 is rn, = p , . K where p, =
1.007 x lo3 kg mP3.'
The molecular weight (M,) of L-77 is = 0.620, so
the number, N , of L-77 molecules in the total volume (VJ
is given by:
N = (m,/M,). N ,
where N, = Avogadro's number = 6.022 x loz3mol-'.
These N molecules will be spread throughout the volume
I/I. Thus, each molecule can be taken to occupy on
average a cubical 'cell' of side I and volume P where:
1= (y/N)'j3
=( . MJp,. K. N,)"'
= (M,/p, N , . C)"3
= (0.620/1.007 x lo3 x 6,022 x loz3x 2 x
= 1.72 x rn
Thus, approximately 400 (i.e. 6.8 x 10-6/1.72 x
molecules of L-77 could be aligned within the thickness
of the droplet film. Therefore, the film could be
considered to be of macroscopic thickness.
3.5.2 Change in bulk concentration due to spreading
In order to investigate the unexpectedly low uptake of
triclopyr-triethylamine at the 1.0 ml litre-' L-77 con-
centration, the effective change in concentration of the
bulk solution within the droplet was calculated for that
concentration, using results of the fluorescence and
radiotracer studies. The following data were employed in
the calculation:
1. The volume of formulation applied to the leaf,
V = 0 . 2 5 ~ 1 (for each of the four replicate
droplets applied).
2. The mean droplet spread or area ( A ) of 0 . 2 5 ~ 1
drops was found to be 37.0 mm2 = 37 x m*.15
3. The density of L-77, p, = 1.007 x lo3 kg m-3.9
4. The area per molecule in tbe monolayer at the
liquid-vapour interface is 70 Az = 70 x lo-'' m2per
molecule).'
The latter area can be deduced from the limiting slope of
the log concentration-surface tension plot (see Fig. 2)
and the Gibb's equation. However, a deficiency in points
R . D.Buick, G . D . Buchan, R . J . Field
below the break point of Fig. 2 created difficulties in
accurately determining this Limiting slope in the present
study, and the value of 70 A2 per molecule determined
elsewhere for an organosilicone similar to L-77 was
used.'
As the droplet spread, an adsorbed monolayer would
have been formed at the solid (leaf)-liquid interface. For
a polyethylene-water interface, the area is approximately
50 A* per m o l e ~ u l eIn
. ~ the absence of a measured value
for adsorption at the leaf-liquid interface, the value for
polyethylene, which, as a hydrocarbon polymer, should
have similar physical properties to cuticular wax, was
used. A wax is an ester formed from a long-chain alcohol
and a long-chain acid, and thus has hydrocarbon
structure along most of its length. The surface adsorp-
tions (kg m-*) of L-77 at the liquid-vapour interface (slJ
and at the solid-liquid interface (s,J were then deter-
mined using:
Surface adsorption (s) =
Mass of one L-77 molecule
Area per molecule at the interface
The surface adsorption at the liquid-vapour interface is:
1.0296x
s, =
70 x 10-2"
= 1.47 x kg m-z
The corresponding surface adsorption of L-77 at the leaf
surface is
1.0296 x
SS1 =
50 x lo-'''
= 2-06 x kg m-'
The final step is to estimate the reduction in the
concentration of L-77 within the droplet. As the droplet
spreads, adsorption into both the interfaces would
produce an 'adsorptive dilution' effect in the bulk
solution. The bulk concentration, C (in this case
1.0 ml litre-') would reduce to C' estimated using the
following equation :
x (1.47 + 2.06) x
= 10-3- 37 x
1.007 x lo3 x 2 3 x 10-lo
= 10-3(1 - o m )
= 0.000481
= 0.48 ml litre-'
Thus, in this case, the bulk concentration is more than
halved. Note that the value of A used is the apparent
foliar spread of the droplet. The actual contact area
could be considerably larger, due to leaf surface
Surface tension in absorption of herbicide via leaf stomata
microtopography, which would accentuate the above
adsorptive dilution.
4 DISCUSSION
Volatilisation losses of triclopyr have been found to be
1ow,22,23 with amine formulations generally exhibiting
smaller losses than ester formulation^.^^ As a result, the
risk of triclopyr losses from field bean leaves was not
considered significant in these uptake studies.
Fluorescence (Table 1) and radiotracer (Table 2)
studies revealed that greatest absorption of triclopyr
occurred when L-77 was included in the solution at 2.5
and 5.0 ml litre-'. The strong association of absorbed
material with stomatal pore regions at these two
concentrations further supported the mode of action of
L-77 involving the promotion of stomatal infiltra-
t i ~ n . ' ' * ' ~ ' 'In
~ contrast, the control solution (no sur-
factant) was absorbed only by glandular trichomes,
surface dust or lesions. The 1.0 ml litre-l L-77 treatment
also appeared to enter trichomes and lesions, but
absorption via stomatal pore regions was limited and
only apparent after 1 h, despite the fact that the surface
tension corresponding to this bulk concentration was
essentially the same as for concentrations of 2.5 and
5.0ml litre-' (Fig. 1). The relatively low uptake of
['4C]triclopyr-triethylamine at the 1.0 ml litre-' L-77
concentration (Table 2) also suggested that a limited
amount of active ingredient was able to enter the leaf,
particularly after 1 h. Although the 2.5 and 5-0 ml litre-'
concentrations of L-77 seemed to be optimum for
triclopyr-triethylamine absorption, evidence exists that
L-77 at 5.0ml litre-' may have a negative effect on
translocation processes.", 18,25"3
A significant reduction in the surface tension of the
triclopyr-triethylamine solution was observed when very
low concentrations of the organosilicone surfactant,
L-77, were added (Fig. 1). A plateau minimum surface
tension was reached at approximately 22 mN m-l. A
dramatic reduction in surface tension is typically obser-
ved when surfactants are added to solution^^-^ but the
magnitude of reduction tends to be greatest for organo-
silicone surfactants.'. l8
3'
Some surfactants induce quite significant changes in
behaviour when concentrations reach or exceed the
CMC6 The formation of micelles in a solution causes a
sharp change in many physical proper tie^,^ of which
surface tension is just one, and may affect the biological
activity of the formulation.6 Figure 2 displays a sudden
break-point or inflexion in surface tension as surfactant
concentration is increased, occurring approximately at
2.0 ml litre-' L-77 for both the amine and water solutions.
Goddard and Padmanabhang estimated the CMC for a
similar organosilicone surfactant in water to be around
this concentration.
233
In the case of organosilicone surfactants, micelle
formation has been found to be limited, with the number
of surfactant molecules in a micelle being as few as 5 to
lo.',' This contrasts with conventional organic surfact-
ants which typically have aggregation numbers ranging
from 20 to 140.7In fact, an organosilicone surfactant,
like L-77, may not form regular micelles at all.26As a
consequence, micellisation may be of minimal import-
ance for L-77 mixtures in terms of biological activity of
the overall formulation.
Surface tension and contact angle results were used to
find a critical surface tension value (ycritical) for the field
bean leaf and formulations used. Contact angles and
surface tension have become frequently used measure-
ments to predict surface wetting and spreading on plant
surfaces.17.2 0 , 2 7 , 2 8 ycriticalis the surface tension at which
the cosine of the contact angle (0,)is unity, i.e.
0,= Oo.20 For the adaxial leaf surface of field bean
with triclopyr-triethylamine and L-77 solutions, the
critical value range was 19.5-22.9 mN m-l (mean of
21.1 mNm-') (Fig. 3). Schonherr and Bukovac' found
ycritical = 25-30 mN m-' for the lower leaf surfaces of 2.
purpusii while Stevens and BukovacZ9found 29 mN m-'
for adaxial maize leaves. The variation in ycritical values
could be due to a number of reasons. Firstly, different
plant species will differ in chemical composition and
spatial arrangement of wax c o n ~ t i t u e n t s . ~Stevens ~~~~~~'
and BukovacZ9discuss how the value of 29mNm-'
seemed high for a very hydrophobic surface like maize
leaves, but they suggested that water repellency of these
leaves is enhanced by epicuticular wax platelet^.'^ Abbott
et aL3' discuss how surface wax roughness greatly
influences ycritiCal values. Furthermore, the kinds of
surfactant solution being studied are important deter-
minants of the critical surface tension value. The above
r e s e a r c h e r ~ studied
~ ~ ~ ~ different solutions which were
different again to the triclopyr solutions used here. A
lower yCritiCal value would be expected if the triclopyr
formulations dissolved the surface waxes of field bean31
which was observed with scanning electron micrographs
where the bean leaf surface appeared to be disrupted or
broken down by triclopyr form~lations.'~ Another aspect
to consider in determining yCriticalvalues is the hydrogen-
bonding properties of liquids, since this has been found
important in the prediction of spreading3'
Zisman plots typically form a rectilinear relation
between the cosine of the contact angle and the surface
tension for a solution series.20s28Determination of the
critical surface tension of liquids from such plots has
proved useful in describing the spreading behaviour of
various liquids on a surface. Critical values have also
been related to the constitution of various low-energy
surfaces. As found with field bean, a critical surface
tension (yCritiCal)of 19.5-22.9 mN m-' on a hydrocarbon
surface is a low value. Shafrin and Zisman" suggest that
these values are typically found with surfaces composed
of closely packed methyl groups.
234
Although this rectilinear relationship is true for most
solutions on most surfaces, the band of data may exhibit
concave curvature for values above 50 mN m-' on some
low-energy surfaces.2oThis was observed with adaxial
leaf surfaces of field bean and was consistent with the
observation that the quadratic curve best described the
relationship (Fig. 3). Up to 45-50 mN m-l the points on
the Zisman plot formed a linear band, but above this
surface tension, the relationship tended to curve up-
wards. It has been shown that such curvature results
from weak hydrogen bonds forming between the molec-
ules of liquid and those in the solid s ~ r f a c e . However,
~'
it should be noted that in this case the quadratic curve
fitted to the cosine @,-surface tension plot was strongly
influenced by only one data point above 40 mN m-'.
Three of the points were below 40 mN m-', improving
the precision of the critical surface tension value
calculated from this part of the curve.
Schonherr and Bukovac' established that stornatal
entry of liquids depends on three main factors: the
surface tension of the liquid, the contact angles of the
liquid on the plant surface, and the stornatal pore wall
morphology. According to their model for stomatal
entry, the surface tension must be lower than the critical
surface tension value for that plant and liquid.
Triclopyr-triethylamine solutions containing 1.0, 2.5 or
5.0 ml litre-' L-77 all appeared to have surface tension
values at or below the critical value for field bean and this
aqueous system, so would be expected to enter stornatal
pores. Transmission electron microscopy found field
bean's stornatal pore walls had angles which would
provide positive pressure differences with the three
triclopyr-triethylamine/L-77 solutions, thus permitting
instantaneous stornatal infiltration.l8 However, the
fluorescence results did not display appreciable levels
of stornatal entry with the solution containing
1.0 ml litre-' L-77. The two higher concentrations (2.5
and 5.0 ml litre-') certainly did facilitate stornatal entry,
so cursory reasoning suggests that surface tension is
unable to fully explain the mechanism by which L-77
enhanced herbicide performance. A lack of correlation
between measured surface tension and performance
of formulations has been observed by several
researchers.6*'6il7
The above investigation into surfactant behaviour in
solution has helped to explain this observation. Sur-
factant molecules tend to aggregate at ~urfaces,~~'' thus
tending to reduce their effective concentration within the
bulk solution. Estimates of this change in bulk con-
centration, and therefore surface tension, following
droplet spreading were determined by considering details
of the surface adsorption.
Initially it was established that the film after spreading
had 'macroscopic' thickness, t , since t was found to be
much greater than the average distance between sur-
factant molecules in solution. The film of aqueous
solution between surfactant monolayers behaves as
R . D.Buick, G.D.Buchan, R. J. Field
normal bulk aqueous solution, and the surface tension
data for macroscopic droplets (Figs 1 and 2) can be used
in investigations of film behaviour.
A deficiency in points below the break point of Fig. 2
created difficulties in accurately determining the limiting
slope and hence the area per moleculeoof L-77, using
Gibb's equation. Thus, the value of 70 A' per molecule,
determined by Goddard and Ananthapadmanabhan' for
an organosilicone similar to L-77, was used to calculate
a surface adsorption of 1-47x kg m-' for L-77 at the
solution-air interface.
Similarly, as the droplet spread, an adsorbed mono-
layer was formed at the solid (leaf)-liquid interface.
Measurements by Goddard and Padmanabhan' in-
dicated that for a polyethylene-water interface tke
adsorption monolayer packed to approximately 50 A'
per molecule. Assuming that this value was appropriate
for adsorption at the leaf-solution interface, the cor-
responding adsorption of L-77 at the plant surface was
found to be 2.06 x kg m-'.
As the droplet spread, adsorption into both the
liquid-vapour and surface-liquid interfaces produced an
'adsorptive dilution' effect in the bulk solution. The
above estimates suggest that a droplet of 1.0 ml litre-'
L-77 solution would actually have had an effective bulk
concentration after spreading of about 0.48 ml litre-', or
even less if the actual droplet contact area exceeded the
apparent area used in the calculation. This is in the most
sensitive region of the surface tension-concentration
curve (Fig. 1) with an apparent surface tension value
similar to the 0.5 ml litre-' solution. The resulting
decrease in bulk concentration may have been sufficient
to raise surface tension to a value above the critical value
(ycritica,), thus inhibiting stornatal entry and herbicide
absorption via this pathway. For 2.5 and 5.0 ml litre-'
solutions, a reduction in effective concentration within
the drop could cause a shift to the left along the
concentration axis (Fig. 1) but as the onset of the plateau
occurred well before the 2.5 ml litre-' value, effective
surface tension would have been substantially unaltered,
remaining well below yCritiCaI. This would allow stornatal
infiltration with the two higher concentrations of
surfactant, according to Schonherr and Bukovac's
model.' In practical terms, herbicide solutions may need
to have a sufficiently high concentration of L-77 (i.e. well
above the inflexion point in the surface tension-log
concentration plot). If the concentration is, for material
economy, selected too close to this point, stomatal
infiltration may be rendered ineffective.
Estimating the adsorptive dilution effect has helped to
explain why the 1.0 ml litre-' L-77 solution was unable to
induce absorption in the same way as the 2.5 and
5.0 ml litre-' solutions. However, these calculations
assume that micelle formation did not significantly
alter the behaviour of surfactant solutions. Although
organosilicone surfactants tend not to form regular and
large micelle~,'~'~ it is not known whether the occurrence
Surface tension in absorption of herbicide via leaf stomata
of even limited micellisation with L-77 in triclopyr-
triethylamine would actually affect surfactant-enhanced
uptake of the herbicide via stomata. Micellisation may
affect the calculations made above, although the small
size of organosilicone micelles (if they form them at all)
and the fact that calculations were made for a
1.0 ml litre-' solution (which would contain relatively
few micelles compared to higher concentrations) should
minimise effects. This area requires further research.
The time taken for absorption was an important
factor. Absorption of triclopyr after 1 min was always
less than after 60 min, particularly for the two higher
concentrations of surfactant. The radiolabelled studies
further confirmed the optimum L-77 concentration as
being above 2.5 ml litre-' for achieving maximum
triclopyr uptake (Table 2). Significant differences be-
tween uptake at these higher concentrations compared to
the 0 and 1.0 ml litre-' concentrations were more obvious
after 1 h than after 6 h. This suggested that most of the
effect of L-77 on triclopyr uptake may occur within the
first hour. Stevens et al.I4 found L-77 exerted most of its
effect on deoxyglucose uptake via field bean stomata
within 10 minutes.
The results presented here provide further evidence to
support the involvement of stornatal infiltration by
triclopyr solutions when the appropriate concentration
of an organosilicone surfactant, such as L-77, is present.
Detailed studies of ycriticis,, surface tension, and the
behaviour of L-77 in solution have revealed a previously
unrecognised phenomenon, referred to here as an
'adsorptive dilution' effect. This phenomenon is a
potential explanation for the limited stornatal infiltration
observed with 1.0 ml litre-l L-77, in spite of this
solution's surface tension apparently being at or below
the critical surface tension value for the field bean
adaxial leaf surface. The first-order calculation suggests
that effective concentrations and surface tension within
droplets may be a useful way to account for stornatal
infiltration of herbicide formulations containing L-77.
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