Journal of Experimental Agriculture International

19(1): 1-22, 2017; Article no.JEAI.36359

ISSN: 2457-0591
(Past name: American Journal of Experimental Agriculture, Past ISSN: 2231-0606)

Changes of Photosynthetic and Antioxidant Activity

of Phaseolus vulgaris to Potassium
Virgílio Gavicho Uarrota1,2*, Cristiane Segatto1, Camila Pereira Barbosa1,
Daicon Godeski Moreira1, Deivid L. V. Stefen1, Emanuel Mattos1,
Gustavo Viana Junkes1, Camila Corrêa1, Marcelo Eduardo Tormem1,
Maira Maier Bisato1, Cileide Maria Medeiros Coelho2 and Clovis Arruda Souza1
1
Laboratory of Crop Plants, Department of Agronomy, Agroveterinary Science Center, Postgraduate
Program in Plant Production, University of the State of Santa Catarina, Luiz de Camões Avenue 2090,
88520-000, Lages, Santa Catarina, Brazil.
2
Laboratory of Seed Analysis, Department of Agronomy, Agroveterinary Science Center,
Postgraduate Program in Plant Production, University of the State of Santa Catarina, Luiz de Camões
Avenue 2090, 88520-000, Lages, Santa Catarina, Brazil.

Authors’ contributions

This work was carried out in collaboration between all authors. Author VGU designed the experiment,
evaluated, performed the data mining in R software and drafted the paper together with the last
author. All other authors contributed equally in data collection, analysis and discussion.

Article Information

DOI: 10.9734/JEAI/2017/36359
Editor(s):
(1) Biljana Bojovic, Assistant Professor, Faculty of Science, Institute of Biology and Ecology, University of Kragujevac,
Republic of Serbia.
Reviewers:
(1) Ramazan Dogan, Türkiye.
(2) Öner Canavar, Adnan Menderes University, Turkey.
Complete Peer review History: http://www.sciencedomain.org/review-history/22169

Received 25th August 2017

th
Accepted 9 November 2017
Original Research Article Published 6th December 2017

ABSTRACT

Greenhouse experiment was conducted with two cultivars of common beans and four
concentrations of potassium (1, 2, 10 and 20 millimolar –mM ) were supplied to the soil pots aiming
to access variations in plant antioxidant defense systems (secondary metabolites and enzymatic
mechanisms), growth parameters and leaf gas-exchange measurements. The total fresh leaf mass
increased in both cultivars from 1 to 10 mM of potassium and the dry matter content decreased from
1 to 20 mM. Root volume significantly increased in Uirapuru cultivar. Potassium increased
chlorophyll A in both in the range of 1 to 10 Mm. Catalase activity and carotenoids increased only in
_____________________________________________________________________________________________________

*Corresponding author: E-mail: [email protected];

 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

Dama cultivar. Potassium increased the photosynthetic activity in cultivar-dependent manner.

Cultivar ‘Dama’ showed higher intrinsic water use efficiency, NDVI index, chlorophyll contents, total
leaf mass and higher catalase activity. If the selection of cultivar under those traits is aimed, ‘Dama’
cultivar is a candidate and presented a linear stomatal conductance model.

Keywords: Common beans; potassium nutrition; photosynthesis; stomatal conductance models;

growth parameters; antioxidant defense mechanisms; phenolics; normalized vegetative
index; catalase.

1. INTRODUCTION dry weight [10]. K provides regulatory control

over different plant processes such as
Common bean (Phaseolus vulgaris L.) is the transpiration, starch synthesis, sucrose
most important grain legume in human diets. It translocation, respiration and lipid synthesis and
provides protein, complex carbohydrates, and has a profound effect on the profile and
valuable micronutrients for more than 300 million distribution of the primary metabolites in plant
people in the tropics. Is the second most tissues [9]. Changes in metabolite concentrations
important source of calories after maize in induced by K are multiple and include K
Eastern and Southern Africa providing 32% of dependence of metabolic enzymes,
energy supply and the fourth in tropical photosynthesis and long distance
America where it is also the third most important transport [9]. The primary metabolites such as
source of calories after maize and cassava and soluble sugars particularly reducing sugars,
constitutes an important source of household organic acids and amino acids tend to
income for small scale farmers [1,2]. The increase in K deficient plants [9]. Experiment
common beans are thought to provide conducted by Rashid [10] found that K
up to 65% of protein [3] and currently estimated fertilization increased K content but reduced N,
to be one of the most important legumes Si, free sugar and soluble protein contents in the
worldwide [4,5]. Beside the great importance, plant tissues which resulted in the minimum
there are many factors that affect the high reduction of relative water content (RWC) in rice
performance of the plant potential and plants.
photosynthesis.
Gautam and co-workers [11] also reported the
Potassium (K+) is the most abundant inorganic beneficial effect of potassium application in
+ improving the submergence tolerance of rice. All
cation in plant cells. K is vital for plant growth.
Together with nitrogen (N) and phosphorous (P), the cultivars evaluated showed inhibition of
+ photosynthesis, and this was accompanied with
K belongs to the top three elements, the
availability of which strongly determines crop decrease in stomatal conductance, chlorophyll
yield. Undeniably, the application of mineral and carbohydrate contents. The activity of
NPK-fertilizers was an essential cornerstone of antioxidants was found to be significantly higher.
the green revolution in the last century [6]. Potassium application improved the survival
Limited resources [6] are now forcing us to mainly because of maintenance of
investigate and better understand how the plants’ carbohydrates, chlorophyll and contributing to
+ less lodging and leaf senescence. Furthermore,
K demand can be satisfied with the minimum of
fertilizer application [6]. The indispensable K application resulted in inhibition of lipid
ground for providing answers to this complex peroxidation and increase in catalase and
question is the understanding of the exact roles peroxidase activities [11]. Potassium at higher
of K+ in plants [6]. Yao & Naeth [7] levels was more beneficial in terms of improving
reported changes in physiological growth stages survival, photosynthesis and plant growth [11].
in barley from emergence and below Experiment conducted in peanut cultivars by
ground biomass in response to potassium Chakraborty [12] showed that external K
application. application resulted in improved salinity tolerance
in terms of plant water status, biomass produced
In plant metabolic processes, potassium under stress, osmotic adjustment and better ionic
application has been reported to inhibit the balance. A study conducted by Zain & Ismail [13]
uptake of cesium [8]. As reported by Rashid [9] in rice aiming to understand the effects of
potassium (K) has a critical role in plant potassium rates (80, 120 and 160 kg/ha) and
physiology and comprises up to 10% of plants types (KCl and K2SO4) on leaf gas exchange,

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growth and biochemical changes under cyclic Aiming to better understanding the role of
water stress concluded that increases in potassium application in photosynthesis,
fertilization rates increased proline production, agronomic attributes, antioxidant behavior and
catalase, malondialdehyde (MDA) and secondary metabolism of common beans,
transpiration rate. Higher potassium rates greenhouse experiment (pot experiment) was
reduced water stress effects by inducing high conducted with two contrasting cultivars (white-
transpiration which increased nutrient uptake to “carioca” and black colored tegument) .
repair the damaged tissues and reduce oxidative
stress. Potassium in KCl form increased net 2. MATERIALS AND METHODS
assimilation rate –NAR. Potassium has been
also reported to play different roles such as 2.1 Plant Material and Soil Preparation
biophysical and biochemical roles, improve
tolerance to biotic and abiotic stress (salinity, Two contrasting commercial common bean
drought, ammonium concentrations, cold, frost, cultivars were selected for this experiment (white
light), reduce oxidative stress load of chilling colored grains-carioca “TAA DAMA”, hereinafter
stressed plants, role in photosynthesis, designed as Dama and black colored grains
movement of plant organs, improve pathogen “IPR88 UIRAPURU”, designed as Uirapuru, see
resistance, maintain osmotic potential of sieve Fig. 1). Dama cultivar is tolerant to rains even
tubes, cell turgor and phloem loading and during harvesting time; it has growth cycle of 85
transport [14-17]. to 95 days, with plant height around 50 cm.
Its growth habit is indeterminate (type III) and
K constitutes about 2.1–2.3% of the earth’s crust the shape of the grain is oblong. It has been
and thus is the seventh or eighth most abundant argued to be moderately resistant to rust, angular
element [14]. Therefore, soil K reserves are stain, mildew and bacteriosis and resistant to
generally large [18]. However, large agricultural common mosaic. Has long duration of light grain
areas of the world are reported to be deficient in color for more than one year; higher productive
K availability. Soils inherently low in K are often ceiling, extensive adaptation and higher sieve
sandy, waterlogged, saline, or acidic. yield [19]. Uirapuru was selected because
Additionally, in intensive agricultural production presents excellent culinary qualities, is
systems, K has become a limiting element, in moderately tolerant to water stress [20], has a
particular in coarse-textured or organic soils. In broad adaptation, with erect stems and branches
many cases, lower fertilizer K application in the which favors direct mechanical harvesting.
context of unbalanced fertilization may result in a Flowers and reaches maturity at 43 and 86 days
significant depletion of available soil K reserves, after germination respectively [21]. Resistant to
and thus in decreased soil fertility [14]. Beside common mosaic virus, rust and powdery
extensive literature of potassium in other crops, a mildew and tolerant to high temperatures [21].
better understanding on how potassium affect Sandy soil in the pots was firstly washed
the photosynthetic and antioxidant is of during03 times (24 hours each) with distilled-
crucial importance to better manage the common deionized water in other to remove any existence
beans. of minerals before seeding.

Fig. 1. (A) Cultivar TAA DAMA and (B)-IPR88 UIRAPURU

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2.2 Greenhouse Experiment and Treat- 2.4 Normalized Difference Vegetative

ments Index (NDVI)

The experiment followed a randomized block As an important indicator of the status of

design, with three blocks. Each block consisted chlorophylls, photosynthetic capacity and energy
of 10 treatments (2 genotypes –“Dama” and absorption by plant canopy, NDVI was measured
“Uirapuru”, hereinafter designed as “UIRA” using analyzer (PlantPen NDVI 300, Photon
and four levels of potassium- 0, 1, 2, 10 and 20 System Instruments, Brazil).
mM applied in a form of potassium chloride
-1
(KCl), molecular weight of 74.548 g mol and the 2.5 Leaf Gas Exchange Data
KCl mass required for each concentration
calculated according to the equation below –Eq. Photosynthetic parameters were measured using
A). These 4 KCl concentration were chosen a portable photosynthesis system-IRGA (LI-
taking into account the previously published 6400XT-LI-COR, Biosciences Inc., Lincoln, NE,
manuscript by Nieves-Cordones [15] which state USA). Data of net photosyntensis (Pn), stomatal
levels of (>10mM) as higher concentration for conductance (gs), intercellular carbon dioxide,
uptake in non-selective channels (main pathways transpiration, leaf to air temperature ratio, leaf
for K+ uptake) while 1mM as intermediate saturation vapor, total conductance to carbon
level[10]. So according to previous reports [10], 2 dioxide, intercellular to ambient carbon dioxide,
high and 2 intermediate levels were chosen and intrinsic water use efficiency (WUE), stomatal
considered adequate for pot experiments. limitation value (SL), carboxylation efficiency of
Treatments in each block were represented by 3 Rubisco were accessed. Intrinsic water use
pots (previously filled with soil) with 3 plants in efficiency, stomatal limitation value and
each in a total of 9 plants per treatment/block. carboxylation efficiency (CarbE) were calculated
Bean seeds were surface sterilized by a according to equations 1, 2 and 3 respectively.
commercial fungicide (carbendazim + tiram at a
dose 300 mL/100 kg of seeds) before seeding in  Pn 
the pots and then 5 seeds were seeded and  W U E =  (1)
placed in the greenhouse under partially  g s 
controlled conditions of light intensity,
photoperiod, relative humidity and temperature.  Ci  (2)
Seedlings were irrigated by basic salt of SL=1- 
Murashige and Skoog nutrient solution (2mM  Ca 
potassium-total concentration and applied 1/4
per week) one time per week, during one month.  Pn 
Plants were watered regularly (one to two times  CarbE=  (3)
per week) with distilled water and 30 days after  Ci 
potassium treatment, the evaluations were
started. Where Ci and Ca means the intercellular and
ambient carbon dioxide respectively, and gs the
 m = [ C ] * Mw*V  (a)
stomatal conductance.

Using “plantecophys”-an R package for analysing

Where “m” is the required mass of KCl; [C] is the and modeling leaf gas exchange data [22],
required concentration; Mw is the molecular photosynthesis and stomatal conductance
weight of KCl and V is the total volume of water models were fitted in other to better understand
used (3.6 L). and quantify inter-specific differences in
photosynthesis and model the rapid response to
2.3 Chlorophyll Content changes in environmental drivers such as light,
humidity and temperature.
Chlorophyll (Chla, Chlb and total) in the
greenhouse was determined using chlorophyll 2.6 Dry Matter, Root Length, Root Volume
electronic analyzer (CLOROFILOG CFL 1030, and Total Leaf Mass
FALKER, serial 0520, Brazil). For that, 18 leaves
in each treatment per block were measured in a These parameters were evaluated at the final of
total of 54 leaves per treatment in all experiment. the experiment, when plants reached the R5-R6

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

phenological stage according to Fernández [23]. 2.0 mL total volume, 200 µL of extract were first
Dry matter of leaves was measured by weighing mixed with 100 µL of FCR reagent after adding
the fresh and dry weight (Forced air oven, 72 1.40 mL distilled water and the contents were
hours, 60°C) of sample and represented as kept at room temperature for 10 minutes. Later,
proportion of dry and wet sample weight (%), 300 µL of Na2CO3 aqueous solution (20%) were
total fresh leaf mass by weighing all leaves added and incubated for 1 hour. The absorbance
collected per treatment and represented in was measured at 765 nm using a
grams, root length using a rule and root spectrophotometer. Total phenolics content was
volume by the water volume dislocated by the expressed as µg of gallic acid equivalents/g of
roots. dry extract (µg GAE/g) using a standard curve
(0-1000 µg/mL) of gallic acid [28].
2.7 Antioxidant Enzymatic System
(Catalase-CAT and Ascorbate UV-visible (UV-vis) scanning (200-800 nm) using
Peroxidase-APX) spectrophotometer of phenolic extracts was also
performed. Spectra were normalized, baseline
Enzymes were extracted from 1 g of leaf tissue corrected, subjected to feature selection at the
using a mortar and pestle with 5 ml extraction region of interest and non-supervised
buffer containing 0.2 M K-phosphate buffer pH7, multivariate analysis (hierarchical cluster
20 mM DTT, 0.100 g PVP, 25 µL triton, 50 mM analysis-HCA and principal component analysis-
PMSF and 5 mM EDTA. The homogenate was PCA).
centrifuged (3000 rpm/30 minutes) and the
supernatant fraction was used to assay catalase, Total flavonoid content of plant extract was
proteins and ascorbate peroxidase [24]. Protein determined using aluminum chloride colorimetric
content was determined using Bradford reagent method (Woisky & Salatino [29] and revised by
[25] with bovine serum albumin as a standard. Chang [30]) and standard solutions (0-1000
Catalase activity was assayed in a reaction µg/mL of quercetin in 80% methanol). For that, 1
mixture of 2 mL of 50 mM pH7 phosphate buffer mL extract solution was mixed with 0.5 mL 95%
containing hydrogen peroxide (25 µL) and 20 µL ethanol (v/v), 0.1 mL 1 M potassium acetate, 0.1
of enzyme extract. The decomposition of mL aluminum chloride solution (10% AlCl3), and
hydrogen peroxide was followed at 240 nm (ε=36 0.8 mL distilled water to a total volume of
-1 -1
mM cm ). Total APX activity was measured by 2.5 mL. The mixture was well mixed and
monitoring the decline in absorbance at 290 nm, incubated at room temperature for 30
as ascorbate (ε= 2.8 mM-1cm-1) was oxidized, for minutes versus reagent blank containing water
2 min using the method of Nakano and Asada instead of sample. Quercetin was used as the
[26]. The assay medium consisted of 600 µL of standard for the quantification of total flavonoid.
50 mM K-PO4 buffer (pH 7.0) with ascorbate, 600 Results were expressed as milligrams of
µL of buffer with hydrogen peroxide and 300 µL quercetin equivalent per gram of dry weight (mg
of sample extract. APX activity was expressed in QE/g).
-1 -1
mM ascorbate.min mg of proteins.
2.9 Data Mining and Statistics
2.8 Non-enzymatic Antioxidant System
(Carotenoids, Total Phenolics and Data were summarized and subjected to
normality (Shapiro-Wilk) and homogeneity
Flavonoids) and UV-Visible Scanning
(Levene) tests and analysis of variance (Two-
of Phenolic Extracts way ANOVA). Tukey HSD (p<0.05) test was
used where differences were found. Multivariate
Carotenoids were determined by the method statistical techniques were also used in other to
described by Lichtenthaler & Buschman [27] find important variables related to potassium in
using cold acetone as a solvent and calculated common beans. All analyses were performed in
by Lambert-Beer equations (4-6) respectively. R software [31] using scripts produced by our
group. A report in html format is provided as
C
 carot ( µ g / ml ) = (1000A
470 −1.90Ca − 63.14Cb ) / 214 (4)

supplementary data for data analysis
reproducibility. Leaf Gas exchange models were
fitted using Plantecophys package [22].
The total phenolic contents of were determined Equations were made in Mathtype software.
by Folin-Ciocalteau reagent (FCR) method. For a

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3. RESULTS AND DISCUSSION of cultivar, the dry matter decreased gradually

(1.5 fold and 1.4 fold) with increases of
3.1 Growth Parameters (Total Fresh Leaf potassium levels in ‘Dama’ and ‘Uirapuru’
Mass, Dry Matter, Root Length and respectively from 1 to 20mM of potassium
Root Volume) (Fig 2B). The ‘Uirapuru’ cultivar presented higher
dry matter (16.7% fold) than ‘Dama’ (P<0.05).
Results of growth parameters are summarized in Tukey HSD test (p<0.05) showed significant
the Fig. 2(A-D). Two-way ANOVA of the total differences between levels 1 and 2 with 10 and
fresh leaf mass showed differences between 20 in ‘Dama’ and between (2 and 10) with (1 and
potassium levels in Dama cultivar. 10mM of 20) in ’Uirapuru’ (see supplementary data for
potassium nutrition provided major total fresh leaf details). Similar results were reported by Taibi
mass (p<0.05) and 1mM the minor value of leaf [33] testing the effect of salt stress (NaCl) in
mass. All other levels (0, 2, 20 mM) were Phaseoulus vulgaris. In their research shoot and
statistically similar (Fig. 2A). Significant statistical root biomass decreased with increases of
differences were found between potassium at 10 salinity. K induces phenotypic dry matter
Mm and other levels (p<0.05). Increases in allocation plasticity and was reported to be
potassium promoted the total leaf mass of responsible of carbon allocation to osmotic
“Dama” cultivar until level 10. For “Uirapuru” adjustment [34]. Root length and volume showed
cultivar, non-significant differences were found a direct relationship between potassium levels.
(Fig. 2A). Previous results by Yaghub [32] also When K concentration increased until 10 mM, the
reported improvement of leaf mass by K root length and volume (Figs. 2C and 2D)
application. Contrarily, K deficiency was found to increased in cultivar dependent manner. In
decrease the leaf number, leaf area and ‘Dama’ cultivar increases were observed until
reproductive dry weight [10]. These changes level 10 of potassium and in ’Uirapuru’ until 2mM
explain our results. Potassium deficiency affects (Figs 2C-D). Besides, root length did not differed
the carbon-nitrogen balance. Improved plant statistically in ‘Dama’ cultivar (p<0.05). Similar
survival and reduction of shoot elongation were trends were also observed for root volume (see
also reported to be affected by K addition [11]. supplementary data). In Sulla carnosa, a forage
legume the growth of vegetative organs was
The dry matter of leaves (Fig 2B) differed decreased by 50% by K deficiency being
between cultivars and potassium levels and was the stems more affected than roots and leaves
affected adversely by potassium. Independently [35].

DAMA
UIRAPURU
40

A
30

ab a
Total leaf mass (g)

ab
a a
20

b
a

a
ab a
10
0

0 1 2 10 20

Potassium levels (mM)

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

DAMA
UIRAPURU

40
a
B

ab
ab

a
a b
30
Dry matter (%)

ab
ab

b
20

c
10
0

0 1 2 10 20

Potassium levels (mM)

DAMA
UIRAPURU
40

C
30

a
a
a
Root length (cm)

a
a ab
ab
b
20

c
10

b
0

0 1 2 10 20

Potassium levels (mM)

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DAMA
UIRAPURU

40
D

30
Root volume (cm3)

20

a
ab ab
10

a a ab

a
b
0

0 1 2 10 20

Potassium levels (mM)

Fig. 2. Total leaf mass (A), dry matter (B), root length (C) and root volume (D) of the two
cultivars subjected to potassium levels. Tukey HSD tests are presented in the supplementary
data

When Pearson correlation of all variables studied and increases water uptake efficiency of roots
(p<0.05) was done (Table 1), the total leaf mass from soil mainly due to the role of K in regulating
(TLM), root volume (RV) and root length (RL) cellular osmotic potentials [36] which may explain
were found to be positively correlated with our observed results. Cultivar ‘Dama’ was most
chlorophyll contents (Table 1) and these last two efficient in activating chlorophyll pigments.
also correlated with catalase activity (Table 1).
The dry matter content of leaves was negatively 3.2.2 Catalase activity (CAT)
correlated with carotenoid content and TLM.
CAT in both genotypes increased until 2mM of
3.2 Photosynthetic Pigments, Enzymatic potassium and then decreased gradually (Fig
and Non-enzymatic Antioxidants 3C). At higher levels of potassium, CAT was less
important as enzymatic antioxidant system. The
3.2.1 Chlorophyll A and B activation of CAT was markedly in ‘Dama’
cultivar than ‘Uirapuru’. ‘Dama’ exhibited plants
Chlorophyll A and B increased with potassium with better performance may be due to the
application (Fig. 3A-B) in all cultivars. The activation of many antioxidant mechanisms.
accumulation of chlorophylls was higher in Contrarily, the level of ascorbate peroxidase
‘Dama’ cultivar than ‘Uirapuru’ (p<0.05). (APX) was higher in ‘Uirapuru’, which decreased
Increases in chlorophyll contents may be with potassium levels (Fig 3D). Trace amounts of
attributed to the role of K in plant metabolism. APX were found in ‘Dama’. Results indicated that
Essentiality of K in plant life cycle is evidenced these 2 enzymatic systems may be of lesser
by its role in activating enzymes which are importance in all genotypes at higher rates of
involved in various physiological processes such potassium.
as energy metabolism, starch synthesis, nitrate
reduction, photosynthesis and sugar 3.2.3 Carotenoids
degradation. As a component of plant
cytoplasmatic solution, K reduces loss of water Carotenoids (Fig. 3E) increased with potassium
from leaf surfaces by regulating stomatal closure levels. The rate was higher in ‘Dama’ than

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Table 1. Pearson correlations of all variables. Chla, Chlb and Chlt mean chlorophyll a, b and total respectively. CAT-catalase activity; APX-ascorbate peroxidase activity; CAR- total
carotenoids; FLA-total flavonoids; PHE-total phenolic compounds; TLM-total leaf mass; DM- dry matter; RL-root length; RV- root volume; TRA-transpiration rate; SVTleaf-leaf
saturation vapor; Cond- stomatal conductance; CndTotal -total stomatal conductance; WUE-intrinsic water use efficiency; Ls-stomatal limitation value; Pn-net photosynthesis;
CarbE- carboxylation efficiency of Rubisco; Ci and Ca-intercellular and ambient carbon dioxide respectively; Ti and Ta-intercellular and ambient temperature respectively and NDVI-
normalized difference vegetative index

Chla Chlb Chlt CAT APX CAR FLA PHE TLM DM RL RV Pn Cond Ci TRA Ti/Ta SVTleaf CndTotal Ci/Ca WUE Ls CarbE
Chla
Chlb 0.98* * * *
Chlt 1.00* * * * 0.99* * * *
CAT 0.43 0.32 0.41
APX 0.04 0.06 0.04 -0.34
CAR -0.38 -0.31 -0.36 -0.26 0.10
FLA -0.35 -0.30 -0.34 0.15 -0.54 -0.01
PHE -0.52 -0.52 -0.52 -0.32 -0.13 -0.13 0.08
TLM 0.61 0.63 0.61 0.19 -0.32 0.29 -0.26 -0.41
DM 0.12 0.11 0.12 0.00 0.42 -0.62 -0.23 0.32 -0.46
RL 0.85* * 0.81* * 0.84* * 0.58 0.16 -0.58 -0.21 -0.43 0.23 0.52
RV 0.59 0.53 0.58 0.62 0.07 -0.46 -0.18 -0.35 0.11 0.53 0.86* *
Pn -0.09 0.02 -0.06 -0.46 -0.17 0.37 -0.13 0.31 0.41 0.02 -0.22 -0.08
Cond 0.10 0.22 0.13 -0.33 -0.10 -0.04 0.00 0.17 0.08 0.27 0.05 0.04 0.64*
Ci -0.51 -0.59 -0.53 -0.10 0.21 -0.14 0.14 0.01 -0.62 -0.01 -0.30 -0.19 -0.62 -0.72*
Tra -0.17 -0.05 -0.14 -0.48 -0.18 0.20 0.03 0.22 0.08 0.07 -0.26 -0.15 0.76* 0.93* * * * -0.59
TI/Ta 0.27 0.38 0.30 -0.19 -0.07 -0.19 0.01 0.12 0.09 0.36 0.24 0.15 0.51 0.97* * * * -0.76* 0.81* *
SVTleaf 0.14 0.26 0.17 -0.28 -0.12 -0.05 0.03 0.15 0.11 0.24 0.08 0.03 0.60 0.99* * * * -0.76* 0.90* * * * 0.98* * * *
CndTotal -0.27 -0.16 -0.24 -0.58 -0.13 0.23 -0.04 0.24 0.04 0.08 -0.34 -0.18 0.80* * 0.85* * -0.46 0.98* * * * 0.70* 0.80* *
Ci/Ca -0.54 -0.62 -0.56 -0.11 0.17 -0.09 0.16 0.02 -0.58 -0.07 -0.35 -0.24 -0.59 -0.74* 1.00* * * * -0.59 -0.79* * -0.78* * -0.46
WUE 0.43 0.35 0.41 0.57 -0.04 0.19 -0.22 -0.26 0.55 -0.33 0.29 0.22 -0.23 -0.66* -0.04 -0.69* -0.58 -0.62 -0.71* -0.01
Ls 0.54 0.62 0.56 0.10 -0.18 0.06 -0.14 -0.01 0.58 0.08 0.35 0.24 0.59 0.74* -1.00* * * * 0.59 0.79* * 0.78* * 0.45 -1.00* * * * 0.01
CarbE 0.14 0.27 0.17 -0.34 -0.16 0.09 -0.04 0.22 0.30 0.19 0.03 0.04 0.83* * 0.95* * * * -0.82* * 0.91* * * * 0.90* * * * 0.94* * * * 0.84* * -0.83* * -0.46 0.83* *
NDVI -0.52 -0.51 -0.52 0.08 -0.61 0.45 0.33 0.14 0.09 -0.39 -0.52 -0.19 0.40 0.24 -0.15 0.49 0.08 0.22 0.50 -0.11 -0.22 0.09 0.26
Significance levels: p < .0001 '****'; p < .001 '***', p < .01 '**', p < .05 '*'
**
Chla, Chlb and Chlt mean chlorophyll a, b and total respectively. CAT-catalase activity; APX-ascorbate peroxidase activity; CAR- total carotenoids; FLA-total flavonoids; PHE-total phenolic compounds; TLM-total leaf mass; DM- dry matter; RL-root length; RV-
root volume; TRA-transpiration rate; SVTleaf-leaf saturation vapor; Cond- stomatal conductance; CndTotal -total stomatal conductance; WUE-intrinsic water use efficiency; Ls-stomatal limitation value; Pn-net photosynthesis; CarbE- carboxylation efficiency of
Rubisco; Ci and Ca-intercellular and ambient carbon dioxide respectively; Ti and Ta-intercellular and ambient temperature respectively and NDVI-normalized difference vegetative index.

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

‘Uirapuru’. Carotenoids, being antioxidants, have gradually with potassium levels until 10mM. Live
the potential to detoxify the plants from the green vegetation was higher in ‘Dama’ cultivar
effects of reactive oxygen species. Carotenoids than ‘Uirapuru’ but was statistically non
are known to function as collectors of light significant (p<0.05, Tukey HSD test). The results
energy for photosynthesis and as quenchers of were in accordance with the total biomass
triplet chlorophyll and O2. Moreover, they observed by that Cultivar. NDVI was higher in
dissipate excess energy via the xanthophyll cycle control plants than those at 20mM of potassium
and can act as powerful chloroplast membrane and was positively correlated with carotenoids,
stabilizers [33, 37, 38]. Increases in carotenoid flavonoids, net photosynthesis, stomatal
contents indicate that those compounds play an conductance, leaf saturation vapour and the
important role under increases of potassium carboxilation efficiency and negatively correlated
levels. As reported by Cazzonelli & Pogson [39], with chlorophyll contents (See Table 1). Similar
carotenoids play essential roles in development, results were also reported by Penuelas [44]
photosynthesis, root mycorrhizal interactions and studying physiological changes in nitrogen and
the production of phytohormones, such as water limited sunflower leaves. Positive
abscisic acid and strigolactones. correlations with chlorophyll and secondary
metabolites were also reported previously in
3.2.4 Flavonoids aquatic vegetation [45-46]. NDVI was also
positively correlated with leaf area index [47].
For flavonoid contents (Fig. 3F), increases NDVI is a broadband index (i.e., normalized
were observed only for ‘Dama’ cultivar. differences between the reflectance in the near
Flavonoids protect the plant against ultraviolet infrared and the red regions of the light
radiations, have antimicrobial properties and act spectrum) that is related to green biomass and
as a deterrent for herbivores by limiting has been used to indirectly estimate
assimilation of dietary proteins and inhibiting photosynthetic capacity and net primary
digestive enzymes [40]. Furthermore, flavonoids productivity [48] and assess whether the target
act as scavengers of free radicals such as being observed contains live green vegetation or
reactive oxygen species (ROS), and also prevent not. NDVI correlates well with canopy features
their formation by chelating metals [41-44, 40]. such as biomass, leaf area index (LAI), absorbed
Increases in flavonoids observed in our photosynthetically active radiation, and canopy
research may be related to antioxidant functions photosynthetic capacity, but fails to capture
of those compounds mainly in ‘Dama’ dynamic physiological processes, which may
cultivar which exhibited plants with better occur on fine temporal and spectral scales
performance. [49,48].

3.2.5 Total Phenolics 3.4 UV-visible Scanning of Phenolic

Extracts
Regarding the phenolic content, decreases were
observed until level 2 of potassium and then All sample spectra showed higher absorbance in
increased in Dama culttivar. Contrarily, for the similar wavelength regions (300-500 nm -
‘Uirapuru’, an increase until 2mM and then region of phenolics and carotenoids and 640-
decreased gradually (Fig. 3G). Phenolics were 700nm (see supplementary Fig. 1). Those
observed to play a role only for ‘Dama’ Cultivar at regions were selected for further multivariate
higher levels of potassium. This group of analysis. HCA analysis showed four different
compounds is claimed to have the ability of free- groups. ‘Dama’ cultivar samples with 1 and 10
radical scavenging and are essential to plant mM of potassium separated alone in distinct
physiology and resistance to biotic and abiotic groups. The third group was composed mainly by
stresses. Besides removing free radicals, these ‘Uirapuru’ cultivar samples with 1, 2, 10, control
compounds are capable of chelating metal, and ‘Dama’ with 20 mM. The fourth group was
activating antioxidant enzymes, and inhibiting composed by ‘Dama’ cultivar samples (control, 2
oxidases [41]. mM) and’Uirapuru’with 20 mM of potassium. The
cophenetic correlation of HCA was 88%
3.3 Normalized Difference Vegetation according to Euclidean distance (Fig. 4A).
Index (NDVI)
Principal component analysis showed that
NDVI results are summarized in the Fig. 3H. As it samples of cultivar ‘Dama’ with potassium levels
can be observed, NDVI index increased of 1mM and 10mM separated alone duo to their

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

composition in chlorophyll contents as it can be total variance explained by PCA was 88.3%,
observed by the loadings or Eigen values of PCA being 48.6 and 33.7% for component 1 (PC1)
(Fig. 4B). Other samples were similar due their and 2 (PC2) respectively.
composition in phenolics and carotenoids. The

DAMA
UIRAPURU
40

A
30
Chlorophyll A(ug/g)

a
a
a a
ab
ab ab ab
b
20

b
10
0

0 1 2 10 20

Potassium levels (mM)

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

100
C DAMA
UIRAPURU

80
Catalase activity (mM/min.mg of protein)

60
40

a
20

ab a ab bc

c b ab ab
b
0

0 1 2 10 20

Potassium levels (mM)

100

D DAMA
UIRAPURU
Ascorbate peroxidase activity (mM/min.mg of protein)

80

a
60

a
40

b
20

b
c

c c
b b b
0

0 1 2 10 20

Potassium levels (mM)

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

DAMA

14
E UIRAPURU

12
10
Total carotenoids (ug/g)

8
6

ab
4

a ab a
a b a a
b
2
0

0 1 2 10 20

Potassium levels (mM)

1400

DAMA
F UIRAPURU
a
1200
1000

a
Total flavonoids (ug/g)

a
a
a
800

a a
a
a
a
600
400
200
0

0 1 2 10 20

Potassium levels (mM)

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

500
G DAMA
UIRAPURU

400
300
Total phenolics (ug/g)

a
a a
a a a a
a
200

a
a
100
0

0 1 2 10 20

Potassium levels (mM)

DAMA
14

H UIRAPURU
12
Normalized difference vegetative index

10
8

a
a a a a
6

a
a a
a
a
4
2
0

0 1 2 10 20

Potassium levels (mM)

Fig. 3. Changes in chlorophyll A (A), chlorophyll B (B), catalase (C), ascorbate peroxidase (D),
total carotenoid contents (E), total flavonoids (F), total phenolic contents (G) and NDVI index
(H) with potassium levels for two common beans cultivars. Values are representative of mean
and standard error of the mean

14
 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

A
UIRA10

UIRA1

UIRA2

DAMA20

UIRANEG

DAMA10

DAMANEG

DAMA2

UIRA20

DAMA1

25 20 15 10 5 0
25

B W_680
W_679
W_678
W_677 DAMA1
W_676
W_650
20

W_651
W_652
W_653
W_654
15
PC 2 (33.7%)

10
5

DAMA10
0

DAMA20
UIRA10
UIRA2
UIRA20
W_367
W_373W_374
W_377
W_376 W_356
W_372 UIRA1
DAMA2 W_370
W_369 W_371
-5

DAMANEG W_355
W_353 W_352
W_351
UIRANEG
W_349W_348
W_350 W_346
W_347
W_302
W_341
W_340
W_339
W_338
W_337
W_336
W_335
W_334
W_333
W_332
W_331
W_330
W_329
W_328
W_327
W_326
W_325
W_324
W_323
W_322
W_321
W_344
W_343
W_342
W_317
W_316
W_315
W_320
W_319
W_318
W_314
W_312
W_311
W_313
W_310
W_309
W_308
W_307
W_345
W_306
W_303
W_305
W_304

-20 -10 0 10

PC 1 (48.6%)

Fig. 4. (A) Hierarchical cluster analysis of UV-Visible scanning spectra of all samples (B) Score
plot and loadings of principal component analysis (B)

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

3.5 Leaf Gas-exchange Data ‘Uirapuru’. The leaf to air temperature

and leaf saturation vapor showed slight
Results of leaf gas exchange are summarized in variation in plants treated with different
the Table 2. The net photosynthesis of all potassium levels, except at level 10 where a
cultivars increased gradually with potassium maxima was observed in ‘Uirapuru’ cultivar. The
levels from 1 to 10mM. The rate of total conductance to carbon dioxide showed
photosynthesis was higher in ’Uirapuru’ cultivar similar trend as in stomatal conductance
except for control plants. A slight increase in previously mentioned in the Fig. 5B. The rate of
stomatal conductance was observed for ‘Dama’ intercellular to ambient carbon dioxide decreased
cultivar. Higher rate of stomatal conductance at in all cultivars, but the decreasing rate was
10mM of potassium was observed for ‘Uirapuru’. higher at 10mM of potassium for ‘Uirapuru’
Intercellular carbon dioxide decreased in all cultivar. Cultivar ‘Dama’ showed water use
cultivars but the rate was higher for ‘Uirapuru’ at efficiency and superior stomatal limitation value
level 10 of potassium. The rate of transpiration in except at 10mM of potassium. Higher rubisco
all cultivars increased with potassium until level carboxilation was also observed at 10mM of
of 10mM. The rate of transpiration was higher in potassium in ‘Uirapuru’.

Table 2. Gas-exchange measurements of the two common bean cultivars. Net photosynthesis,
stomatal conductance, intercellular carbon dioxide, Transpiration rate, Leaf to air temperature
ratio, Leaf saturation vapor, Total conductance to carbon dioxide, Intercellular to ambient
carbon dioxide, Intrinsic water use efficiency, Stomatal limitation value and Carboxylation
efficiency of rubisco

Dama cultivar**
Potassium amount 0 1 2 10 20
mean SD mean SD mean SD mean SD mean SD
Pn 6.31 1.6 2.82 2.4 3.32 0.9 5.19 2.6 3.50 2.1
Cond 0.10 0.0 0.04 0.0 0.04 0.0 0.08 0.0 0.04 0.0
Ci 274.05 2.0 299.69 64.4 258.32 25.3 261.44 16.8 249.16 29.6
Trmmol 2.60 0.6 1.04 0.5 1.15 0.2 1.77 1.0 1.07 0.5
Tl/Ta 0.57 0.3 1.29 0.2 1.28 0.1 0.97 0.4 1.24 0.2
SVTleaf 4.02 0.1 3.70 0.2 3.85 0.1 3.75 0.1 3.88 0.2
CndTotal 0.10 0.0 0.04 0.0 0.04 0.0 0.07 0.0 0.04 0.0
Ci/Ca 0.70 0.0 0.75 0.2 0.65 0.1 0.67 0.0 0.63 0.1
WUE 61.52 2.3 53.43 32.6 75.70 16.0 71.97 11.3 82.43 14.8
Ls 0.30 0.0 0.25 0.1 0.35 0.1 0.33 0.0 0.37 0.1
CarbE 0.02 0.0 0.01 0.0 0.01 0.0 0.02 0.0 0.01 0.0
Uirapuru cultivar
Pn 3.25 0.8 3.77 3.1 3.76 2.6 6.67 3.2 5.29 3.0
Cond 0.09 0.0 0.07 0.0 0.08 0.1 0.70 0.7 0.07 0.0
Ci 311.20 26.1 302.73 21.2 301.69 6.1 205.81 129.2 275.08 55.6
Trmmol 2.31 1.0 1.33 0.4 1.66 1.3 4.79 0.9 1.66 0.7
Tl/Ta 0.73 0.4 1.15 0.2 0.97 0.6 6.18 8.8 1.01 0.4
SVTleaf 4.01 0.1 3.60 0.4 3.48 0.5 10.97 10.2 3.69 0.3
CndTotal 0.09 0.0 0.06 0.0 0.07 0.1 0.15 0.1 0.07 0.0
Ci/Ca 0.79 0.1 0.76 0.1 0.75 0.0 0.51 0.4 0.70 0.1
WUE 41.96 16.0 51.12 14.5 55.01 9.7 30.97 22.6 67.57 16.3
Ls 0.21 0.1 0.24 0.1 0.25 0.0 0.49 0.4 0.31 0.1
CarbE 0.01 0.0 0.01 0.0 0.01 0.0 0.05 0.0 0.02 0.0
** Net photosynthesis (Pn). stomatal conductance (Cond). intercellular carbon dioxide (Ci). Transpiration rate
(Trmmol). Leaf to air temperature ratio (Tl/Ta). Leaf saturation vapor (SVTleaf). Total conductance to carbon
dioxide (CndTotal). Intercellular to ambient carbon dioxide (Ci/Ca). Intrinsic water use efficiency (WUE). Stomatal
limitation value and Carboxylation efficiency of rubisco (CarbE)

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

0.094
(a)

0.093
)
−1
Modelled gs ( mol m s
−2

0.092
0.091

Uirapuru Cultivar
Control
1mM
0.090

2mM
10mM
20mM

0.075 0.080 0.085 0.090 0.095 0.100 0.105

−2 −1
measured gs (mol m s )
0.1045

(b)
0.1040
)

0.1035
−1
Modelled gs (mol m s
−2

0.1030
0.1025
0.1020

Dama Cultivar
Control
1mM
2mM
10mM
20mM

0.100 0.102 0.104 0.106

−2 −1
measured gs (mol m s )

Fig. 5. Modeled (with the model of Medlyn et al., 2011) versus measured stomatal conductance
(gs) for the two cultivars studied under different potassium levels. (A) for ‘Uirapuru’ and (B) for
‘Dama’ cultivars

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

Photosynthetic assimilation of carbon is a assimilation of carbon dioxide responds to

defining feature of plant kingdom. The fixation of potassium levels. Furthermore, stomatal
large amount of carbon dioxide supports the movements provide the leaf with the opportunity
synthesis of carbohydrates, which make up the to change both the partial pressure of carbon
bulk of plant biomass. Measurements of carbon dioxide at sites of carboxylation and the rate of
assimilation rates are therefore crucial duo to transpiration [51,54-56]. For ‘Dama’ cultivar (Fig.
their impact on the plant metabolism, growth and 5B) at all levels of potassium, the fitted models
reproductive success [50]. As it can be observed showed a linear relation, but such trend was not
in our results, potassium nutrition in the range of observed for ‘Uirapuru’ (Fig 5A).
(1-10 mM) promoted higher carbon assimilation.
The decrease of intercellular carbon dioxide may Data of all variables studied were also correlated
be attributed to the higher transpiration rate and (Pearson correlation, p<0.05) aiming to find an
assimilation activity observed. Cultivar ‘Dama’ association between them. As it can be observed
was most efficient in water use and showed in the Table 1, a positive correlation of root
lower stomatal limitation value. Higher carbon length with chlorophylls was found. Significant
assimilation rates in ‘Uirapuru’ cultivar may be positive association was also observed in net
attributed to the elevated stomatal limitation photosynthesis with stomatal conductance and
value. The carboxylation of rubisco was more carboxylation efficiency (Table 1). The stomatal
pronounced at 10 mM of potassium in all conductance was negatively correlated with the
cultivars. It is important to note that the presence intercellular carbon dioxide and intrinsic water
of finite stomatal resistance to carbon dioxide use efficiency. The transpiration rate was
cause carbon dioxide at sites of carboxylation to positively correlated with leaf temperature, leaf
be less than that in the atmosphere, thereby saturation vapor and stomatal conductance.
reducing the rate of carbon dioxide assimilation Contrarily, a negative association of NDVI index
somewhat below its potential. Nevertheless, with chlorophyll contents was observed (see
stomata usually impose the largest resistance to Table 1). A positive correlation between the
diffusion [51]. Lower stomatal values in ‘Dama’ stomatal limitation value with the carboxylation
cultivar may have contributed to lower carbon efficiency of rubisco was also found.
dioxide assimilation rates observed in this
research. Previous report has indicated that K When all dataset was subjected to principal
deficiency promotes inhibition of enzymatic- component analysis (PCA) aiming to
photochemical processes [52]. Results indicate dimensionality reduction and find important
lower chlorophyll content and altered Rubisco variables, similarities and dissimilarities between
activity as probable causes of photosynthetic the samples (Fig. 6), an interesting result was
impairment. Potassium deficiency was found to found, as it can be observed in the score plot and
diminish photoprotection mechanisms due to loadings of PCA. Control samples grouped
reduced photosynthetic and photorespiratory together showing that there are similar and
capacity [52]. composed the first group in PCA. Such an
observation was also found for those samples
In the past, stomatal responses have generally where 1mM of potassium was applied. Samples
been considered in relation to single of ’Uirapuru’ cultivar with 2 and 20 mM of
environmental variables in part because the potassium grouped together. A clear separation
interactions between factors have appeared was observed from ’Uirapuru’ at 10mM of
difficult to quantify in a simple way. A linear potassium and the control samples. Samples of
correlation between stomatal conductance (gs) ‘Dama’ cultivar at 2, 10 and 20 grouped also
and carbon dioxide assimilation rate has been together duo to their catalase activity and
reported when photon fluence was varied and intrinsic water efficiency presented. Intercellular
when the photosynthetic capacity of leaves was carbon dioxide and the ratio of intercellular to
altered by growth conditions, provided C02, air ambient carbon dioxide where the parameters
humidity and leaf temperature were constant that most contributed to sample clustering of
[53]. In this research using the measured gs we control plants (see Fig 6). Contrary, ’Uirapuru’
fitted the stomatal conductance models of the samples at 10mM stayed alone duo to phenolic
two cultivars (Fig. 5A-B). A linear (‘Dama’ contents, high level of NDVI index, stomatal
cultivar) and non-linear regression was found for conductance, net photosynthesis, transpiration
‘Uirapuru’ cultivar (Fig 5A-B). As we know, and stomatal limitation value. The total variance
fundamental to any examination of stomatal explained by the model was 64.3%, being 36.6
functioning is the understanding of how rate of and 27.7% for component 1 (PC1) and 2 (PC2)

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

NDVI CndTotal
PHE
DAMA_0 TRA
CAR

4
Pn
UIRAPURU_0 FLA
Ci/Ca
Ci Cond
CarbE
SVTleaf
2
Ti/Ta
PC 2 (27.7%)

UIRAPURU_10
0

DAMA_1
UIRAPURU_1 Ls
UIRAPURU_20
UIRAPURU_2
TLM
-2

DAMA_2
WUE CAT
DAMA_10
RV

DAMA_20 Chlb
RL Chlt
Chla
-4

-2 0 2 4 6 8 10

PC 1 (36.6%)

Fig. 6. Score and loading plot of Principal component analysis of all dataset showing sample
clustering and the main variables correlated. The total variance explained by the two first
components were 36.6% and 27.7% respectively

respectively. Most samples grouped in (PC1- plant biomass, root length and volume and
/PC2-) axis. The control samples grouped in the chlorophyll contents. Increases in carotenoids,
(PC1-/PC2+) axis and ’Uirapuru’ at 10 mM in flavonoids and catalase activity may be
(PC1+/PC2+) axis. correlated to their antioxidant activities and as
scavengers of reactive oxygen species that can
The loading values indicated that sample be produced during the high photosynthetic rate
clustering in PC1 were highly influenced by the and transpiration. Cultivar ‘Dama’ was most
net photosynthesis, stomatal conductance, tolerant to potassium levels and presented a
intercellular carbon dioxide, transpiration, linear relationship of stomatal conductance.
intercellular to air temperature, carboxylation Potassium levels also increased net
efficiency and stomatal limitation value. Those photosynthesis, NDVI indices, carboxylation
clustered in PC2 were most influenced firstly by efficiency and transpiration, but the variation was
the chlorophyll contents, followed by catalase in a cultivar-dependent manner. Intercellular
activity, phenolic contents, root length, root carbon dioxide decreased with increases of
volume, transpiration, water use efficiency and potassium levels. Multivariate tools revealed that
NDVI index. most samples of ‘Dama’ cultivar were most
influenced by CAT and the intrinsic water use
4. CONCLUSIONS efficiency presented than ‘Uirapuru’.

The results of this study prompt us to conclude SUPPLEMENTARY DATA

that there are considerable photosynthetic and
biochemical differences between the cultivars A report in html format is provided for data
studied in their response to potassium nutrition. reproducibility. Detailed statistical analysis not
Potassium range from 1 to 10mM promoted the shown in the manuscript can be found here.

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 Uarrota et al.; JEAI, 19(1): 1-22, 2017; Article no.JEAI.36359

ACKNOWLEDGEMENTS host rice plants. Rice Science. 2016;

23:119-131.
The financial support of CAPES-BRAZIL and 10. Hu W, Coomer T, Loka D, Oosterhuis D,
CNPq-BRAZIL is to be acknowledged. The first Zhou Z. Potassium deficiency affects the
author thanks CAPES for supporting the carbon-nitrogen balance in cotton leaves.
postdoctoral fellowship under the PNPD Plant Physiology and Biochemistry. 2017;
program. 115:408-417.
11. Gautam P, Lal B, Tripathi R, Shahid M,
COMPETING INTERESTS Baig M, Maharana S, Puree C, Nayak A.
Beneficial effects of potassium application
Authors have declared that no competing in improving submergence tolerance of rice
interests exist. (Oryza sativa L.). Environmental and
Experimental Botany. 2016;128:18-30.
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