Vol.4, No.11 A, 19-25 (2013) Agricultural Sciences
Physiological potential of soybean seeds treated
with thiamethoxam and submitted to storage
Lilian Gomes de Moraes Dan1*, Alessandro Lucca Braccini1,
Alberto Leão de Lemos Barroso2, Hugo de Almeida Dan1, Gleberson Guillen Piccinin1,
Juliana Marques Voroniak1
1Department of Agronomy, State University of Maringá, Maringá, Brazil;
*Corresponding Author: liliangmdan@yahoo.com.br
2Department of Agronomy, University of Rio Verde, Rio Verde, Brazil
Received 18 July 2013; revised 18 August 2013; accepted 23 September 2013
Copyright © 2013 Lilian Gomes de Moraes Dan et al. This is an open access article distributed under the Creative Commons Attribu-
tion License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly
Bio-activators such as thiamethoxam have been
used to increase the productive potential of
plants, by means of metabolic and plant mor-
phology modifications. Nevertheless, very little
is known regarding the effectiveness of this in-
secticide when incorporated into lower quality
level soybean seeds, p articularly during storage.
The objective of this work was to assess the in-
fluence of thiamethoxam on the physiological
potential of soybean seeds with different initial
vigor levels during four storage periods. Three
vigor levels of cultivar Anta—low (35%), medium
(70%) and high (85%) vigor—of soybean seeds
were utilized in this experiment. The experi-
ment al de sig n was completely randomized in a 3
× 2 × 4 factorial scheme—3 levels of vigor (low,
medium and high); 2 seed treatments (with and
without thiamethoxam) and 4 storage pe rio ds (0,
30, 60 and 90 days). The analy zed v ariables were:
germination, first germination count, acceler-
ated aging test, seedling length and emergence
speed index. Thiamethoxam-treated soybean
seeds benefit the physiological potential of both
medium and high vigor seeds during storage up
to 30 days. Whereas, the use of thiamethoxam
was shown to be ineffective in improving the
physiological quality of initial low vigor seeds.
Keywords: Glycine max; Bio-Activator; Vigor;
Physiological Quality
Soybean crops comprise both increased productive po-
tential and adaptability in different regions of Brazil due
to recent technological advances in the productive seg-
ment. Among the modern farming techniques, the incor-
poration of products such as fungicides and insecticides
to seeds in order to protect the plants and increase the
productive potential is becoming an increasingly wide-
spread practice in most technified agriculture in ad-
vanced countries [1].
In general, insecticides are assessed for their efficiency
on pest control; however, some products can cause ef-
fects yet unknown, modifying the metabolism and mor-
phology of the plant [2]. The discovery of the molecule
thiamethoxam (3-(2-chloro-thiazol-5-ylmethyl)-1,3,5-
oxadiazinan-4-ylidene-N-(nitro)amine) brought about new
perspectives for agriculture, mainly regarding seeds’
treatment. A number of studies have shown the efficiency
of thiamethoxam as a bio-activator, promoting incre-
ments on germination, emergence, seedling length (roots
and aerial parts) and dry phytomass. In addition, the
product can eventually improve the performance of
plants under stress, besides increasing total protein and
enzyme content levels in plants [3]. Thiamethoxam
proved to be efficient as a bio-activator, essentially for
soybean seeds [4].
According to [5], thiamethoxam positive physiological
effect is related to germination, vigor, enzymatic activity,
nutrient level, height plant, stem diameter, radicular de-
velopment, phytomass, pods’ number per plant, grain
weight and yield, providing an increment, on average of
four bags per hectare of soybean. Increased vigor, pro-
ductivity, leaf and radicular area index, a more uniform
stand, emergence uniformity and better initial develop-
ment of soybean seedlings were observed by [6].
Thiamethoxam used in soybean seed treatment, ac-
celerates germination and induces further development
Copyright © 2013 SciRes. OPEN A CCESS
L. G. de Moraes Dan et al. / Agricultural Sciences 4 (2013) 19-25
of embryonic axis, minimizing the negative effects in
situations such as presence of aluminum, salinity and
water deficit. This acceleration of germination is due to
the stimulation of peroxidase activity, preventing oxida-
tive stress [7], thus, reducing the time of crop establish-
ment in field, and therefore, lessening the harmful effects
of either weed competition or essential nutrients present
in soil [2].
Nevertheless, it is equally important to consider that
the effectiveness of thiamethoxam treatment may depend,
among a number of factors, on seed vigor at the time of
product application. According to [8], the utilization of
biologically active compounds such as bio-activators can
either cease or reduce the impact of adverse factors on
seeds’ quality and performance. Moreover, one of the
greatest limitations of soybean farming is unavailability
of high-vigor seeds at the sowing time [9]. Additionally,
little is known about the bioactive effect of thiameth-
oxam applied to initial low- or medium-vigor soybean
Another factor to be considered is that soybean seeds
usually present reduction in physiological quality during
long term storage [10], even under favorable conditions.
In Brazil, the preservation of soybean seeds’ quality dur-
ing storage is among the largest obstacles for seed indus-
try. According to [11], seed quality maintenance until
sowing time ensures the increased emergence potential
of seedlings in field, resulting in productivity gains. Dur-
ing storage, seed quality can either remain close to the
initial level or reduce to a level which makes the seed
unavailable for sowing. This fact is related to several
circumstances, such as environmental conditions during
seed production, occurrence of pests, oil content in seeds,
water content, mechanical damages over the course of
processing and packaging, temperature and relative hu-
midity of the air during storage [12-14], besides chemical
treatment of seeds using insecticides [15,16].
Thus, thiamethoxam proved to be effective as a bio-
activator on soybean seeds’ treatment, implying the ne-
cessity to assess its effectiveness regarding its use in
lower physiological quality level seeds, particularly dur-
ing storage.
In this scope, this present study had the objective to
assess the influence of thiamethoxam on physiological
potential of soybean seeds with different initial vigor
levels in four periods of storage.
2.1. Experiment Installation and Conduction
This experiment was conducted at the State University
of Maringá (UEM), Maringá, Paraná State, and assess-
ments on physiological quality of seeds were carried out
at the Laboratory of Seed Technology at the Applied Ag-
riculture Research Nucleus (NUPAGRI), Center for Ap-
plied Agricultural Research of UEM.
Soybean seeds of cultivar Anta 82 RR from Adriana’s
Seeds—maturity group 7.4 and semi-determinate growth
habit—were used in this study. Tetrazolium test was used
for this cultivar [17] to select lots with three levels of ini-
tial vigor: low (35%), medium (70%) and high (85%).
The experimental design was completely randomized in
a 3 × 2 × 4 factorial scheme—3 levels of vigor (low, me-
dium and high); 2 seed treatments (with and without
thiamethoxam) and 4 storage periods (0, 30, 60 and 90
Chemical treatment of soybean seeds was carried out
using thiamethoxam (Cruiser® 350 FS), at a dose of 2 mL
per kg of seeds.
Syrup (product + distilled water) was applied using a
graduated pipette at the bottom of a transparent plastic
bag and spread across its walls up to 15 cm high. The
volume of syrup was 0.3 L 100 kg1 of seeds. The wit-
ness received distilled water on the same volume of
syrup with the insecticide. The quality of treated seeds
was assessed at the following times: 0, 30, 60 and 90
days after soybean seed treatment.
2.2. Assessed Features
The following tests were carried out both at the labo-
ratory of seeds and greenhouse using trays containing
washed and sterile sand, and then utilized to assess the
physiological quality of soybean seeds.
2.2.1. Germination Test
Four subsamples of 50 seeds were used for each lot
and replicate use; they were then placed for germination
between three-leaf germination paper imbibed in distilled
water in a ratio of 2.5 times as much the paper dry mass.
They were then rolled up and placed in a Mangelsdorf
germinator kept at a constant temperature of 25˚C.
Evaluations on the percentage of normal seedlings were
carried out both on the fifth (first count) and eighth (final
count) days, according to the established criteria of the
Seed Analysis Rules [18]. First count was considered as
an indicative of vigor and final count as the total seeds
germination percentage.
2.2.2. Accelerated Aging T est
Plastic boxes (gerbox) provided with horizontal
stainless steel screen fixed at the median position were
used in this experiment. 40 mL of distilled water (in or-
der to get approximately 100% of Relative Humidity)
was added to the bottom of each gerbox, and seeds were
uniformly distributed on the screen in order to cover its
surface, forming a single layer. Subsequently, the boxes,
together with the seeds, were sealed and stored in a Wa-
ter Jacket Incubator and kept at a constant temperature of
Copyright © 2013 SciRes. OPEN A CCESS
L. G. de Moraes Dan et al. / Agricultural Sciences 4 (2013) 19-25 21
41˚C for 48 hours [19]. After this period, seeds were
taken for germination under the same conditions de-
scribed previously on germination test.
2.2.3. Emergence speed index
It was carried out from sowing in trays containg 8 kg
of washed and sterile sand in 4 replicates of 50 seeds for
each sample. Test was conducted at the greenhouse and
emerged seedlings were counted daily between the on-
sets (five days after installation) of emergence until the
numerical stabilization of the counts (ten days after in-
stallation). Emergence speed index calculation was ac-
complished in accordance with [20], using the formula
proposed by [21].
2.2.4. Seedling Length
Five samples of 20 seeds from each treatment were
distributed in germination paper rolls imbibed with dis-
tilled water in a 2.5:1 ratio (mL distilled water per dry
paper mass in grams) and kept into a germination cham-
ber at 25˚C for five days [20]. A line was longitudinally
traced in the upper third upon the humidified paper towel,
where seeds were placed pointing the micropyle down-
ward. Seedlings length which was considered normal [18]
was determined at the end of the fifth day using a milli-
meter-gauged ruler.
2.3. Statistical Analysis
Variables which characterize the physiological quality
of seeds were all submitted to variance analysis
(ANOVA) using the Statistical Analysis System-SISVAR.
Regression analysis was utilized to verify polynomial
adjustment for dependent variables according to storage
periods, at 5% probability [22].
3.1. Germination Assessment
Significant interaction was shown for both treatment
of seeds and periods of storage (Figure 1) by assessing
the germination test results. For high and medium-vigor
seeds, submitted or not to thiamethoxam treatment, re-
ductions in germination performance over storage were
similar. Nevertheless, taking into account the study ac-
complished by [7], thiamethoxam, used for seed treat-
ment accelerated soybean germination, whose effect was
pronounced when twice-fold dose of that recommended
for the crop was utilized (4 mL·kg1 of seed).
Furthermore, at 90-day storage, germination percent-
ages were higher to 80% in both seed treatments assessed,
thus being within the trading-accepted standards of soy-
bean seeds in Brazil [23]. In thiamethoxam-treated soy-
bean seeds any germination and vigor differences were
found [24], as well as [25] who observed no germination
Figure 1. Polynomial regression for soybean seeds germina-
tion percentage with different levels of initial vigor, treated (T)
and not treated (NT) with thiamethoxam in four periods of
effects on bean seeds.
The lack of thiamethoxam positive effects on germina-
tive performance in this current work may be on the ba-
sis of the recommended dose used for the crop (2
mL· k g 1 of seed). However, the choice for using the
recommended dose in this study was mainly not only due
to economic viability but also to environmental and eco-
logical aspects related to chemical dose increments.
For low vigor thiamethoxam-treated seeds (Figure 1),
47% germination was observed in the onset of storage,
whereas for non-treated seeds, germination level was
38%. This fact shows the positive action of thiameth-
oxam treatment in low-vigor seeds, when not stored.
Nevertheless, the effect of storage is negative for both
treated seeds and for those which received no chemical
treatment. These results are in accordance with [16] who
observed that soybean seeds treated or not with thia-
methoxam showed germination reduction over a 45-day
3.2. First Count of Germination Test
Significant interaction between seed treatment and
storage period (Figure 2) was observed at the polyno-
mial regression assessment for normal seedling percent-
age in first germination count. In high-vigor soybean
seeds, treatment using thiamethoxam and the witness
showed normal seedling percentage of 0.15% and 0.14%,
respectively in each day’s storage. Thus, demonstrating
similarity to that reduction of physiological performance
of initial high-vigor seeds among treatments during stor-
In both medium- and low-vigor seeds (Figure 2), re-
ductions on normal seedling percentage in the first count
were 0.10% and 0.17%, respectively for each day’s stor-
Copyright © 2013 SciRes. OPEN A CCESS
L. G. de Moraes Dan et al. / Agricultural Sciences 4 (2013) 19-25
Figure 2. Polynomial regression for normal seedling percent-
age in the first count of soybean seed germination with dif-
ferent initial vigor levels, treated (T) and not treated (NT) with
thiamethoxam in four periods of storage.
age for thiamethoxam-treated seeds. Whereas, for the
witnesses (no chemical treatment), vigor decrements
were 0.16% (medium vigor) and 0.21% (low vigor) for
each day’s storage. These results support thiamethoxam
beneficial effects on vigor maintenance of soybean seeds
with initial quality ranging from intermediate to low
during storage [26], when working with cotton crops,
concluded that higher increments are reached by using
thiamethoxam in lower physiological quality seeds, par-
ticularly in vigor tests. Thiamethoxam promotes soybean
seed expression related to both enzyme synthesis and
activity by modifying amino acids of phytohormone pre-
cursors, resulting in an increase of germination, vigor
and radicular growth [27].
Nevertheless, at day 90 of storage, only initial high-
vigor seeds showed normal seedling percentage over
80%, despite the reduction observed along with storage
period increment.
3.3. Vigor Assessment in the Accelerate
Aging Test
Results on accelerated aging test (Figure 3) showed
that in initial high-vigor thiamethoxam-treated seeds,
decrement was more intense in normal seedling percent-
age during storage, ranging from 89.8% at the beginning
of storage to 57.4% after 90-day storage. On the other
hand, assessment carried out right after thiamethoxam
application (period zero) in seeds, percentages of 89%
for vigor and 83% for treatment without the insecticide
were observed. These results show that the positive ef-
fects on initial high-vigor soybean seeds is limited to
short term storage when submitted to stress conditions in
the accelerated aging test.
Figure 3. Polynomial regression for normal seedling percent-
age after accelerated aging test of soybean seeds with different
initial vigor levels, treated (T) and not treated (NT) with thia-
methoxam in four periods of storage.
In medium-vigor seeds, thiamethoxam incorporation
via seed treatment presented positive action during stor-
age because angular coefficient indicated 0.16% reduc-
tion on seed vigor for each day’s storage. In contrast, for
the witness, seed vigor decrease after accelerated aging
test was 0.19% in each day of storage (Figure 3).
Accelerated aging test is one of the most used for
physiological potential assessment of several species
[28]. This test has as key principle lead to a considerable
increase on seed deterioration when exposed to high lev-
els of temperature and relative air humidity, which are
considered preponderant environmental factors for both
intensity and speed deterioration. It is notorious that
stress conditions induce reactive oxygen species (ROS),
which damage biomolecules, leading to cellular death.
However, there are antioxidative enzymes such as su-
peroxide dismutase and peroxidase which are responsible
for ROS elimination. According to [7], thiamethoxam
speeds up seed germination under stress conditions by
eliminating the ROS generated. Thiamethoxam moves
through plant cells and activates various physiological
reactions, such as the expression of functional proteins
related to defense mechanisms of plants against stress
factors, for instance, water deficit, high temperatures,
toxic effects, just to name a few [4]. Thus, the influence
of thiamethoxam on these defense mechanisms and on
antioxidative enzyme activities may have played a role
against adverse conditions of high temperature and in-
creased relative humidity imposed by the aging test, and
therefore, having attenuated the physiological potential
decrement of soybean seeds during storage.
In initial low-vigor seeds (Figure 3), the use of thia-
methoxam showed no positive effects on the improve-
ment or maintenance of seed vigor during storage for 90
days, once angular coefficient presented the same reduc-
Copyright © 2013 SciRes. OPEN A CCESS
L. G. de Moraes Dan et al. / Agricultural Sciences 4 (2013) 19-25 23
tion in both seed treatments (treated and not treated).
3.4. Emergence Speed Index Assessment
In the results of emergence speed index (ESI) (Figure
4), reductions observed upon this variable were also sig-
nificant as storage period increased in both seed treat-
ments. ESI reductions observed over the course of stor-
age, initial high- and low-vigor seeds highlight thia-
methoxam negative effect on soybean seedling emer-
gence speed derived from treated seeds and submitted to
storage. In contrast, this insecticide treatment in medium
vigor seeds showed similar effect when compared to
those of the witnesses on the ESI performance, present-
ing 0.05 reduction unit each day in which treated seeds
were stored. These results are conflicting to those found
by [4], who concluded that soybean seeds treated with
thiamethoxam showed faster germination due to the ac-
tion of the product that stimulates enzymatic activity,
providing a more uniform and better initial seedling de-
3.5. Seedling Length Assessment
Soybean seedlings height (Figure 5) showed signifi-
cant reduction (p < 0.05) as storage period increased in
both seed treatments. In initial high- and medium-vigor
thiamethoxam-treated seeds, reductions on seedling
lengths in the order of 0.08 and 0.04, respectively, were
observed in each day of storage. In the witnesses, reduc-
tions of 0.04 and 0.03 cm were observed in each day of
storage for high- and medium-vigor seeds.
Despite the highest reduction on this response variable
during storage is determined in thiamethoxam treatment,
seedling length was 4.94 cm taller than the witness for
initial high-vigor seeds at the storage period onset (pe-
riod zero).
In medium-vigor seeds, the maximum efficiency of the
product, for this variable, enabled seedlings height to in-
crease up to 3.0 cm, comparatively to seeds with no
product applications. Thus, that the bioactive effect of
thiamethoxam in initial high- and medium vigor-seeds is
limited to a 30-day storage period is evidenced. The
thiamethoxam tendency to increase seedling length cor-
roborates with the effect observed by [24] in soybean
After a 30-day period, the natural aging process of
seeds, which occurs during storage and culminates in
physiological quality loss, negatively interferes in the
thiamethoxam action in order to increase seedling growth.
This fact can also be observed on results of seedlings
derived from initial low-vigor seeds, treated or not with
thiamethoxam, which showed the same reduction during
storage. Thus, the absence of thiamethoxam positive in-
fluence on seedling length from low-vigor seeds may be
Figure 4. Polynomial regression for the emergence speed in-
dex (ESI) of seedlings derived from soybean seeds with dif-
ferent initial vigor levels, treated (T) and not treated (NT) with
thiamethoxam in four periods of storage.
Figure 5. Polynomial regression for seedling length (cm) de-
rived from soybean seeds with different initial vigor levels,
treated (T) or not treated (NT) with thiamethoxam in four pe-
riods of storage.
due to the advanced deterioration degree which these
seeds presented, indicating that this insecticide shows no
effect as a bio-activator in low quality seeds.
An overall review of data obtained with soybean cul-
tivar Anta suggests that, for most assessments, greater
increments were reached using thiamethoxam in higher
physiological quality seeds. In addition, physiological
quality of soybean seeds was favored by thiamethoxam
treatment, providing more expressive effects right after
its application up to a 30-day storage period for treated
seeds. These characteristics of thiamethoxam, along with
the use of increased genetic and physiological quality
seeds maximize seed production capacity.
Copyright © 2013 SciRes. OPEN A CCESS
L. G. de Moraes Dan et al. / Agricultural Sciences 4 (2013) 19-25
Soybean seed treatment using thiamethoxam favors
the physiological potential of high- and medium-vigor
seeds during 30-day storage. In initial low-vigor seeds,
the use of thiamethoxam is inefficient in improving
physiological quality.
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