Journal of Plant Nutrition

ISSN: 0190-4167 (Print) 1532-4087 (Online) Journal homepage: http://www.tandfonline.com/loi/lpla20

Effects of NaHCO3 on photosynthetic

characteristics, and iron and sodium transfer in
pomegranate

H. R. Karimi & F. Eini Tari

To cite this article: H. R. Karimi & F. Eini Tari (2017) Effects of NaHCO3 on photosynthetic
characteristics, and iron and sodium transfer in pomegranate, Journal of Plant Nutrition, 40:1,
11-22, DOI: 10.1080/01904167.2016.1161770

To link to this article: https://doi.org/10.1080/01904167.2016.1161770

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Published online: 28 Apr 2016.

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 JOURNAL OF PLANT NUTRITION
2017, VOL. 40, NO. 1, 11–22
http://dx.doi.org/10.1080/01904167.2016.1161770

Effects of NaHCO3 on photosynthetic characteristics, and iron and

sodium transfer in pomegranate
H. R. Karimi and F. Eini Tari
Department of Horticultural Sciences, Faculty of Agriculture, University of Vali-e-Asr, Rafsanjan, Iran

ABSTRACT ARTICLE HISTORY

In order to study effects of sodium bicarbonate (NaHCO3) stress in irrigation Received 8 January 2014
Downloaded by [RMIT University Library] at 06:05 08 December 2017

water on photosynthetic characteristics and iron (Fe) and sodium (NaC) Accepted 17 November 2014
translocation content of pomegranate plants, a factorial experiment was KEYWORDS
conducted based on completely randomized design with three cultivars of Chlorophyll index; Fv/Fm;
pomegranate (“Gorch-e-Dadashi,” “Zagh-e-Yazdi,” and “Ghermez-e-Aliaghai”) pomegranate; stress
and four concentrations of NaHCO3 (0, 5, 10, and 15 mM), with three
replications. The results of plant analysis indicated that NaHCO3 affected
chlorophyll index, Fv/Fm, and performance index (PI) in upper and lower
leaves of shoots and also the translocation of NaC and Fe. The results also
showed that Fe translocation from root to shoot reduced at 15 mM level of
NaHCO3. The highest Na translocation and the lowest Fe translocation were
observed in Zagh-e-Yazdi and Ghermez-e-Aliaghai cultivars, respectively. The
ratio of sodium/potassium (NaC/KC) in stems was higher than that in roots
and leaves, and the observed chlorophyll content of upper leaves was also
lower than that of lower leaves. Based on the measured parameters Gorch-e-
Dadashi cultivar showed less relative sensitivity than other cultivars to
NaHCO3 of irrigation water through maintaining the lower NaC transport to
the shoots, and improvement of Fe transport to shoots.

Introduction
Pomegranate (Punica granatum L.) is native to Iran and grown extensively in arid and semi-arid
regions of the world) Sarkhosh et al., 2006). Currently, most of pomegranate cultivation is in arid
and semi-arid regions of the world, where soil salinity, alkalinity, and water stress are the main limit-
ing factors in appropriate yield production. This problem not only limits growth, but also causes
decrease in quantity and quality of fruits. In these regions, rain is not adequate to wash lime and
consequently discharge it from soil profile. The presence of only lime in soil is not problematic in
nutrient elements absorption, but heavy irrigation, soil compacting, and every cause that decreases
aeration of the soil lead to an increase in carbon dioxide concentration. Consequently, lime produces
bicarbonate that has a buffering property. Buffering property of bicarbonate prevents the decrease of
soil pH in rhizosphere (Shahabi and Malakouti, 2005). The presence of bicarbonate is one of the rea-
sons for alkalification of water irrigation. When alkalinity of soil solution increases with accompa-
nying high pH, availability of some microelements and also P decreases (Jones, 2001). Zhang et al.
(2012) recorded that NaHCO3 induces an intracellular increase in pH and the sodium (NaC) concen-
tration in root cells increases with the increasing concentration of NaHCO3, and consequently
NaHCO3 reduces the regulatory effect of selective absorption of NaC. They also showed that
NaHCO3 induced a significantly higher HC efflux than sodium chloride (NaCl) after addition to the

CONTACT H. R. Karimi h_karimi1019@yahoo.com Department of Horticultural Sciences, Faculty of Agriculture, University of

Vali-e-Asr, Rafsanjan, Iran.
© 2017 Taylor & Francis Group, LLC
 12 H. R. KARIMI AND F. E. TARI

test solution in tobacco. Bicarbonate is one of the main factors causing Fe chlorosis in strategy I
plants (Mengel et al., 1984). Usually in alkaline soil, adequate iron (Fe) translocates from roots to
shoots, but reduction of Fe3C to Fe2C in leaves is disturbed by high pH of apoplastic space indirectly
(Amri and Shahsavar, 2009). In a study carried out on uptake and translocation of Fe59 in grapevine
rootstocks, Nikolic et al. (2000) concluded that bicarbonate-induced Fe chlorosis was caused by an
inhibition of Fe uptake and translocation due to inhibition of Fe3C reduction by root cells. These pro-
cesses were less inhibited in the chlorosis-resistant rootstocks that hint to genotypical differences in
Fe acquisition by roots at high bicarbonate levels. Fe is immobile in plants; therefore, deficiency
symptoms appear in young leaves. Interveinal chlorosis is the main symptom associated with Fe defi-
ciency, followed by reduction in shoot and root growth, complete foliar chlorosis, defoliation, shoot
dieback, and under severe conditions tree mortality. By acidifying cell sap, using sulfuric acid
(H2SO4) in irrigation water and/or by direct usage of H2SO4 in soils, nutrients availability in soil and
plant can be increased. Decreasing the acidity of whole soil is not possible and economic in many
conditions (Bindra, 1983). Thus, irrigation by acidified water has a large effect on chemical compo-
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nent of soils and changes the compounds of reactive phases and available nutrients for plants (Van
et al., 2008). In addition, if the reason for high bicarbonate levels is poor soil aeration, these levels
will decrease by soil drainage and successive culturing. Higher plants, due to fixing stability in soil,
do not have feasibility of escaping from abiotic environmental stresses. Therefore, the plants that sur-
vive become equipped with tolerance, resistance, and compatibility (Rao et al., 2006). One of the eco-
logical solutions of these problems is use of efficient cultivars and rootstocks selection techniques.
Pomegranate belongs to strategy I plants that under Fe deficiency are able to increase Fe reductase
activity and net release of protons and organic compounds in roots (Brancadoro et al., 1995; Jimenez
et al., 2007). There is little information about water management in pomegranate orchards (Intri-
gliolo et al., 2011). In spite of many researches on the effects of different concentrations of NaHCO3
on different plants, no study has been conducted regarding the effects of different concentrations of
this anion in alkaline soil and irrigation water on pomegranate. Therefore, this study attempted to
examine the effects of different concentrations of NaHCO3 in water irrigation on photosynthetic
characteristics and some elements distribution in some cultivars of pomegranate.

Materials and methods

Cuttings and performance of treatments
The experiment was conducted in the research greenhouse of Horticultural Department, Faculty of Agri-
culture, Vali-e-Asr University of Rafsanjan, Kerman, Iran for 1 year. The cuttings of pomegranates were
obtained from the pomegranate collection in agriculture, Vali-e-Asr University of Rafsanjan. The cuttings
were rooted and transplanted to plastic pot containing 5 kg of soil. Sandy loam soil with pH of 7.8 in water
and electrical conductivity (ECe) of 1.2 dS/m was used for this experiment. The experiment was arranged
as a factorial in the framework of a completely randomized design with two factors including three pome-
granate cultivars [“Gorj-e-Dadashi” (GD); “Zagh-e-Yazdi” (ZY); “Ghermez-e-Ali Aghai” (GHA)] and
four levels of NaHCO3 (0, 5, 10, and 15 mM), with three replications. Rooted cuttings were exposed to
NaHCO3 treatment for 70 days. Considering field capacity, every pot was irrigated with 500 mL of water,
every 5 days. The temperature fluctuation was between 24 and 33
C during the experiment.

Photosynthetic parameter measurement

Chlorophyll index of leaf (SPAD index) [Fv/Fm and performance index (PI)] was monitored periodi-
cally during the experiment on three different areas of the three expanded leaves (lower leaves) and
three young leaves (upper leaves) by using a portable chlorophyll meter (SPAD MINOLTA 502, Osaka,
Japan) and a portable pocket Plant Efficiency Analyzer (PEA, Hansatech Instruments Ltd., Norfolk,
UK), respectively. The plant leaves were dark-adapted for 20 min by fixing special tags on each leaf
blade before fluorescence measurement.
 JOURNAL OF PLANT NUTRITION 13

Element analysis
At the end of experiment, pomegranate plants were cut at the soil surface and roots were washed to
remove the soil. Plants were divided into roots, stems, upper leaves, and lower leaves. Samples of leaves,
shoots, and roots were turned to ash in a muffle oven at 550 § 25
C. The resulting white ash was then
dissolved in 10 mL of 2 N hydrochloric acid (HCl) and the volume was adjusted to 100 mL for determi-
nation of NaC, KC, and Fe concentrations. The NaC content was measured according to the Chapman
method (1961). The Fe concentration was measured using an atomic absorption instrument (Model
3110) and NaC and KC concentrations were measured using a flame photometer instrument (Model
JENWAY). The translocation rate was calculated according to the following equation:

AR ðshootÞ D ðdry weight of shoot £ concentrationÞ

AR ðrootÞ D ðdry weight of root £ concentrationÞ
TR D AR ðshootÞ=½AR ðshootÞ C AR ðrootÞ£ 100
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Data analysis
Analysis of variance (ANOVA) was performed using SAS and Minitab softwares. If the effects of the
treatments were significant (p < 0.05), then the means were compared by least significant difference
(LSD) test. In these cases, the standard error of the interaction means (SEM) was calculated.

Results
Chlorophyll index
According to ANOVA, the effects of NaHCO3, cultivar, and interaction of NaHCO3–cultivar on the
chlorophyll index of upper and lower leaves were significant (p < 0.01) (Table 1). Chlorophyll index of
lower leaves of GD and GHA cultivars at 15 mM NaHCO3 decreased significantly, whereas it did not
show a significant effect on ZY cultivar. The decrease percent of chlorophyll index in GD and GHA
cultivars at 15 mM NaHCO3 in comparison to control was 8.76% and 24.63%, respectively. Therefore,
the lowest chlorophyll index of lower leaves belonged to GHA cultivar. Chlorophyll index of upper
leaves of GD and GHA cultivars did not change significantly with NaHCO3, whereas it decreased sig-
nificantly in ZY cultivar in comparison to control. Chlorophyll index of upper leaves was lower than
that of lower leaves; for example, at 15 mM NaHCO3 chlorophyll index of lower leaves GD cultivar
was 43.07, whereas it was 31.89 in upper leaves (Table 2).

Table 1. Analysis of variance of the NaHCO3 and cultivar effect on measured parameters of pomegranate.
Mean square

Lower leaves of shoot Upper leaves of shoot

NaC/KC NaC/KC NaC/KC

Degrees Chlorophyll Chlorophyll Fe trans NaC trans ratio of ratio of ratio
Variable of freedom PI Fv/Fm index PI Fv/Fm index location location leaves stems of roots

Cultivar 2 0.066ns 0.0016 110.63 3.170 0.0005 ns 61.38 26.385ns 33.603 .003 12.377 0.007
NaHCO3 3 .416 0.0015 34.43 1.194 0.0009ns 56.93 140.354 90.928 0.010 3.867 0.006
Cultivar£ 6 0.584 0.0011 41.33 2.199 0.0005 ns 18/77 292.436 32.393 0.007 8.690 0.006
NaHCO3
Error 22 0.117 0.0002 3.69 0.032 0.0003 4.84 9.104 3.370 0.0006 0.127 0.0008
(%)CV 20.673 2.044 3.95 6.970 2.528 6.37 13.937 2.055 14.681 16.940 15.526

and mean significant at 5% and 1% probability levels, respectively.
ns
,non-significant.
 14 H. R. KARIMI AND F. E. TARI

Fv/Fm of lower and upper leaves

According to ANOVA, the Fv/Fm of lower leaves was significantly influenced by NaHCO3, cultivar, and
interaction of NaHCO3 and cultivar (p < 0.01) (Table 1). Fv/Fm of lower leaves decreased in all culti-
vars with increasing bicarbonate levels (Figure 1). The response cultivars were different regarding
NaHCO3 so that Fv/Fm of ZY cultivar decreased from 5 mM NaHCO3, while it occurred at 10 and
15 mM in GD and GHA cultivars, respectively. The highest decrease percent of Fv/Fm of lower leaves
(9.54%) was observed in GD cultivar. The effect of cultivar, NaHCO3, and their interaction on Fv/Fm of
upper leaves was not significant (Table 1).

PI of lower and upper leaves

Effect of NaHCO3 on PI of lower leaves (p < 0.05) and effect of interaction of NaHCO3 and cultivar
were significant (p < 0.01) (Table 1). PI of lower leaves of GD and ZY decreased at 15 mM NaHCO3,
while GHA cultivar was not influenced by NaHCO3 levels (Table 2). PI of upper leaves was signifi-
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cantly affected by cultivar, NaHCO3, and interaction of NaHCO3 and cultivar (p < 0.01) (Table 1). PI
of upper leaves of GD cultivar decreased at 15 mM NaHCO3, despite a significant increase at 5 and
10 mM NaHCO3, in comparison with control. PI of upper leaves of ZY and GHA cultivars decreased
by 29.11% and 60.43%, respectively, as NaHCO3 levels rose. Therefore, the highest decrease belonged
to GHA cultivar (Table 2).

Iron translocation
According to ANOVA, Fe translocation was significantly influenced by NaHCO3 and interaction of
NaHCO3 and cultivar (p < 0.01) (Table 1). As NaHCO3 levels raised, Fe translocation in all culti-
vars, GD, ZY, and GHA, recovered at 5 and 10 mM, respectively (Figure 2). At 15 mM, in compar-
ison with control, Fe translocation in ZY and GHA cultivars decreased significantly, while in GD it
was higher than control. The highest rate of Fe translocation (49.80%) was observed in GD cultivar
(Figure 2).

Sodium translocation
The effect of NaHCO3, cultivar, and interaction of NaHCO3 and cultivar on NaC translocation
(p < 0.01) was significant (Table 1). In all cultivars, rate of NaC translocation increased with increasing
NaHCO3 concentration, and the highest translocation rate was observed in ZY cultivar at 10 and
15 mM NaHCO3 (Figure 3).

NaC/KC ratio in leaves

Effects of cultivar and interaction of NaHCO3 and cultivar on NaC/KC ratio in leaves were significant
(p < 0.01) (Table 1). In ZY and GHA cultivars, NaC/KC ratio in leaves increased significantly com-
pared with control at 10 and 15 mM NaHCO3, so that NaC/KC ratio in leaves of ZY cultivar at 15 mM
was 2.5-fold of control. NaC/K ratio in leaves of GD cultivar decreased from 10 mM NaHCO3 (i.e.,
64.70%) in comparison with control. The GD and GHA cultivars had the lowest and highest NaC/KC
ratios in leaves at 15 mM NaHCO3, respectively (Table 3).

NaC/K ratio in stems

The NaC/KC ratio in stems increased in GD cultivar at 5 and 10 mM NaHCO3 in comparison with
control and after that decreased at 15 mM, whereas it did not show any significant difference compared
with control (Table 3). The NaHCO3 concentrations in irrigation water increased the NaC/KC ratio by
7- and 2.29-fold in stems of ZY and GHA cultivars, respectively, in comparison with control. The
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Table 2. Interactive effect of sodium bicarbonate and cultivar on photosynthetic parameters in lower and upper leaves of pomegranate.

NaHCO3 (mM)

Lower leaves of shoot Upper leaves of shoot

Cultivar 0 5 10 15 0 5 10 15

Chlorophyll index
GD 47.21 § 0.24efy 44.71 § 0.69gf 47.81 § 1.00e 43.07 § 1.78g 35.32 § 1.19bcy 35.37 § 0.75bc 34 § 0.75bc 31.89 § 1.76c
ZY 51.60 § 1.27bcd 51.73 § 2.32b 53.74 § 0.30ab 49.93 § 1.83cde 42.2 § 1.30a 36.04 § 1.48b 35.05 § 1.84bc 34.40 § 1.58bc
GHA 55.54 § 0.68a 48.36 § 0.52de 47.16 § 0.61ef 41.86 § 0.10g 35.09 § 1.25bcy 35.07 § 1.13bc 34.52 § 1.46bc 32.99 § 0.40bc
PI
GD 1.16 § 0.15dy 2.22 § 0.12a 2.12 § 0.12a 1.19 § 0.25d 1.4 § 0.18ey 2.92 § 0.23c 2.36 § 0.11d 1.34 § 0.17e
ZY 1.95 § 0.17ab 1.71 § 0.11a-d 1.33 § 0.13cd 1.28 § 0.23d 3.40 § 0.03ab 3.09 § 0.02c 2.42 § 0.11d 2.41 § 0.06d
GHA 1.87 § 0.27abc 1.96 § 0.34ab 1.59 § 0.20bcd 1.4 § 0.10bcd 3.69 § 0.14ay 3.44 § 0.04ab 3.21 § 0.09bc 1.46 § 0.13e

GD, Gorch-e-Dadashi; GHA, Ghermez-e-Ali Aghai; LSD, least significant difference; ZY, Zagh-e-Yazdi.
Data are mean § SE of three replicates.
y
Means in each row or column with the same letters are not significantly different at 5% level of probability using LSD test.
JOURNAL OF PLANT NUTRITION
15
 16 H. R. KARIMI AND F. E. TARI

Figure 1. Interactive effect of NaHCO3 and cultivar on Fv/Fm in lower leaves of pomegranate. Data are mean § SE of three replicates
Control: Bic 0 mM; Bic 5 mM: 5 mM concentration of NaHCO3; Bic 10 mM: 10 mM concentration of NaHCO3; Bic 15 mM: 15 mM concen-
tration of NaHCO3. GD, Gorch-e-Dadashi; GHA, Ghermez-e-Ali Aghai; ZY, Zagh-e-Yazdi.

lowest and highest NaC/KC ratios in stems at 15 mM NaHCO3 were observed in GD and GHA culti-
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vars, respectively.

Root NaC/KC ratio

The results showed that the effects of cultivar, NaHCO3, and interaction of cultivar and NaHCO3 on
NaC/KC ratio in roots were significant (p < 0.01) (Table 1). The results showed that in GD and ZY cul-
tivars, NaC/KC ratio in roots increased as concentration of NaHCO3 increased to 10 mM and then
decreased to 15 mM. In GHA cultivar, NaC/KC ratio in roots significantly increased at 15 mM
NaHCO3. The highest and lowest NaC/KC ratios in roots at 15 mM NaHCO3were observed in GHA
and GD cultivars, respectively. Comparison of NaC/KC ratio in stems, leaves, and roots showed that
NaC/KC ratio in stems was higher than that in leaves and roots, especially in GHA cultivar (Table 3).

Discussion
According to the results of this study, chlorophyll index changed with respect to the place of leaves on
shoot, so that chlorophyll index of upper leaves was lower than that of lower leaves. Previously, similar
reports were observed in rose (Valdez-Aguilar and Reed, 2007), quince, and pear rootstocks (Prado
and Alcantara-Vara, 2011). As Fe has an important role in chlorophyll synthesis and more than 90%
of Fe in leaf cell is located in chloroplasts (Terry and Abadia, 1986), one of the reasons for decrease in
chlorophyll index is leaf Fe deficiency that leads to disorder in photosynthesis. Furthermore, decrease
in chlorophyll index of upper leaves as compared with that of lower leaves attributes to prevention of
absorption and translocation elements, especially Fe toward young leaves (near the apical bud) and

Figure 2. Interactive effect of NaHCO3 and cultivar on iron translocation rate of pomegranate. Data are mean § SE of three replicates.
Control: Bic 0 mM; Bic 5 mM: 5 mM concentration of NaHCO3; Bic 10 mM: 10 mM concentration of NaHCO3; Bic 15 mM: 15 mM concen-
tration of NaHCO3. GD, Gorch-e-Dadashi; GHA, Ghermez-e-Ali Aghai; ZY, Zagh-e-Yazdi.
 JOURNAL OF PLANT NUTRITION 17

Figure 3. Interactive effect of NaHCO3 and cultivar on sodium translocation rate of pomegranate. Data are mean § SE of three repli-
cates. Control: Bic 0 mM; Bic 5 mM: 5 mM concentration of NaHCO3; Bic 10 mM: 10 mM concentration of NaHCO3; Bic 15 mM: 15 mM
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concentration of NaHCO3. GD, Gorch-e-Dadashi; GHA, Ghermez-e-Ali Aghai; ZY, Zagh-e-Yazdi.

consequently it accumulates in lower leaves and roots (Zuo et al., 2007). These conditions lead to the
reduction of chlorophyll in young leaves, which results in chlorosis. In addition to the mentioned con-
ditions, the following was presented: Iron chlorosis expanded severely in young leaves because they are
a growing part of plant with incomplete xylem structure. Therefore, they would not be able to directly
translocate Fe from roots toward young leaves, without the presence of large amounts of nutrients
(Aref, 2011). This decrease maybe attributed to the formation of proteolytic enzymes such as chloro-
phyllase, which is responsible for the chlorophyll degradation (Sabater and Rodriguez, 1978) as well as
damaging the photosynthetic apparatus (Yasseen, 1983). The results are in agreement with those of
Shahabi et al. (2005) on apple, Bie et al. (2004) on lettuce, Chartzoulakis (1994) on cucumber, and
Banuls and Primo-Millo (1992) on citrus. In chlorosis of leaves arising from alkalinity stress, Fe trans-
location along a vegetative growth course was observed. In this manner, the reduction in chlorophyll
content of leaves (chlorosis symptoms) shifted to the youngest leaves (new flashes), so that previous
flashes regreened again parallel with the formation and changing of these new leaves (new flashes)
(Bybordi and Tabatabaei, 2009). Results of this study confirmed the findings of Nikolic and Romheld
(2002) in grapevine, Pestana et al. (2005) in citrus, and Tagliavini et al. (2000) in apple.
Iron translocation in GD and ZY cultivars did not decrease more in the presence of high NaHCO3 con-
centration (15 mM), although Fe translocation in GD cultivar was low at control treatment. GHA cultivar

Table 3. Interactive effect of NaHCO3 and cultivar on leaves, stems, and roots NaC/KC ratio of pomegranate.
NaHCO3 (mM)

Cultivar 0 5 10 15
C
Leaves Na /K ratio
GD 0.17 § 0.024abcy 0.18 § 0.023ab 0.09 § 0.024de 0.06 § 0.008e
ZY 0.06 § 0.011e 0.07 § 0.008e 0.10 § 0.045de 0.15 § 0.008bc
GHA 0.11 § 0.024cde 0.14 § 0.012bcd 0.24 § 0.024a 0.21 § 0.017a
Stem NaC/K ratio
GD 0.65 § 0.39efy 2.01 § 0.108bc 1.87 § 0.037c 0.72 § 0.415ef
ZY 0.31 § 0.057f 0.81 § 0.12def 2.58 § 0.19b 2.17 § 0.24bc
GHA 1.51 § 0.520de 1 § 0.615 def
2.50 § 0.297b 3.47 § 0.028a
Roots NaC/K ratio
GD 0.19 § 0.018bcdy 0.22 § 0.015bc 0.27 § 0.006a 0.11 § 0.029f
ZY 0.10 § 0.026f 0.19 § 0.017bcde 0.17 § 0.010de 0.15 § 0.008ef
GHA 0.17 § 0.012cde 0.19 § 0.005bcde 0.20 § 0.012bcd 0.23 § 0.013ab

GD, Gorch-e-Dadashi; GHA, Ghermez-e-Ali Aghai; LSD, least significant difference; ZY, Zagh-e-Yazdi.
Data are mean § SE of three replicates.
y
Means in each row or column with the same letters are not significantly different at 5% level of probability using LSD test.
 18 H. R. KARIMI AND F. E. TARI

was not resistant to increasing NaHCO3 concentration. The ability of GD and ZY cultivars to recover can
be due to various reasons. It may be due to excretion of phenolic compounds to lower pH, because strat-
egy I plants respond to Fe deficiency via physiologic and morphologic changes. These responses include
rhizosphere acidification via proton efflux, excretion of phenolic compounds and organic acids for Fe che-
lation, and reduction of Fe3C to Fe2C (Guerinot and Yi, 1994; Hajiboland et al., 2005).
In cucumber plants (Colangelo and Guerinot, 2004), the release of proton, due to its ability to
decrease pH in root apoplast and its effect on neutralizing bicarbonate (HCO3)–, provided better con-
ditions for reducing Fe that is trapped in apoplast of root epidermal cell (Kosegarten et al., 2001) and
had become physiologically inactive under NaHCO3 stress (Jarrahi et al., 2013). This strategy is effec-
tive because with every one unit of drop in pH, Fe3C will be more soluble (Olsen et al., 1981). It is
observed that GD cultivar is more efficient than other two cultivars in Fe translocation from stems to
leaves, considering that the effect of NaHCO3 did not decrease this cultivar Fe translocation rate.
Results of this study confirmed the findings of Zuo et al. (2007) and Rao et al. (1987) on peanut, Cinelli
(1995) on quince, and Byrne and Rouse (1994) on citrus. According to the results of this study, relative
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recovery in response to high HCO3– concentrations can be related to decrease in the amount of
NaHCO3, because of fixation of HCO3– ion with calcium as calcium carbonate (Carrow and Duncan,
2011; Yang et al., 2007).
The highest and the lowest decrease percent of PI in lower leaves belonged to GD and ZY cultivars,
respectively. The highest and the lowest decrease percent of PI in upper leaves was observed in GHA
and GD cultivars, respectively. GD cultivar showed the highest decrease percent of Fv/Fm in lower
leaves of shoot at 15 mM NaHCO3, while ZY cultivar was not affected. In GD cultivar, PI of lower
leaves at 15 mM NaHCO3 recovered and ZY cultivar was affected by NaHCO3at 10 mM. The highest
decrease in percent of PI was observed in ZY cultivar. In agreement with these results, Fv/Fm decreased
in chlorotic leaves induced by high bicarbonate in citrus (Pestana et al., 2005), peach rootstocks
(Molassiotis et al., 2006), cucumber plants grafted onto squash rootstocks (Appenroth et al., 2001),
and pea (Jelali et al., 2011). According to Deng et al. (2010), Fv/Fm remained relatively constant at low
and moderate salinity–alkalinity, and decreased only when salinity–alkalinity concentration was high
enough and PI showed a greater sensitivity than Fv/Fm, which is in agreement with many findings (van
Heerden et al., 2003; Christen et al., 2007).
Many investigations showed that Fv/Fm of plants under moderate stress usually changes little, and the
decline in Fv/Fm ratio under severe stress reflects a reduction in the ability of Photosystem II to reduce the
primary acceptor quinone a (Calatayud and Barreno, 2001). It is usually explained as an increase in num-
ber of the closed PSII reaction centers, which do not participate in electron transport (Toth et al., 2005).
As this was observed in ZY cultivar, it was concluded that Fv/Fm ZY cultivar would be affected when
higher NaHCO3 concentration is tested on it and duration of exposure to high NaHCO3 concentration is
increased. Considering that intervenial chlorosis of the younger leaves is a classical symptom of Fe defi-
ciency induced by alkalinity stress, in this study effect of NaHCO3 on Fv/Fm of upper leaves of shoot was
not significant. Therefore, on the basis of PI, photosynthetic efficiency of GD cultivar was better than that
of other two cultivars, which confirmed the results of studies on Salvia (Kang and van Iersel, 2004).
PI is a multiparametric expression that combines the three main functional steps taking place in
PSII (light energy absorption, excitation energy trapping, and conversion of excitation energy to elec-
tron transport) (Strauss et al., 2006). The present results are in agreement with study by Deng et al.
(2010) on chlorophyll fluorescence and gas exchange responses of maize seedlings to saline–alkaline
stress as PI decreased gradually with increase of alkalinity levels. Considering that lower and upper PI
of ZY cultivar decreased, it seemed that this cultivar showed the lowest photosynthetic apparatus per-
formance. It was expectable, because its chlorophyll content decreased and the effect of Fe deficiency
on chloroplast structure as basic foundation of photosynthetic apparatus is undeniable. The reasons
regarding the conflict of ZY cultivar Fe translocation have been discussed previously. The present
results in chlorophyll fluorescence confirmed that photosynthetic activity was more inhibited under
high alkalinity than that under low and moderate alkalinity. With the increasing alkalinity concentra-
tion, the electric transfer was gradually inhibited and energy dissipation enhanced and finally the com-
prehensive photosynthesis PI and photosynthesis rate decreased (Deng et al., 2010). This decrease is
 JOURNAL OF PLANT NUTRITION 19

related to Fe content of leaves, chlorophyll content, and PI in cultivars, because Fe is necessary for chlo-
rophyll biosynthesis (Pushnik and Miller, 1989).
GD cultivar showed higher tolerance to sodium bicarbonate than ZY and GHA cultivars through
maintaining chlorophyll content and lower NaC transport to the shoots, and improvement of potas-
sium transport to shoot. However, the ability of pomegranate cultivars in prevention of NaC transport
to shoot seems to be dependent and limited to roots. It means if absorption and/or transport of NaC
are controlled at root level, pomegranate cutting would be successful. Naeini et al. (2004) reported that
up to 40 mM of NaC was accumulated in root cells of pomegranate, which translocated to shoot at
higher salinity levels.
In this study, stems showed a higher NaC/KC ratio than leaves and roots, which was likely due to less
C
Na transport to shoots or to older leaves in pomegranate. Under alkali stress, plants frequently accumu-
late large amounts of NaC as a main osmolite to lower the cell water potential (Munns and Tester, 2008).
It is well known that many plant species have a NaC exclusion mechanism that is dependent on exchange
of NaC/HC antiport, which exchanges cytoplasmic NaC with external HC (Munns and Tester 2008). The
exchange activity relies on the transmembrane proton gradient achieved by HC-ATPase. Under alkali
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stress, the lack of external protons might weaken the exchange activity of the NaC/HC antiport on the
root plasma membrane, possibly reducing the exclusion of NaC into the rhizosphere and enhancing in
vivo accumulation of NaC, even to toxic levels that may be the basis of alkali injury (Chen et al., 2011).
Therefore, the main reason is related to more tolerance of GD cultivar, considering that all conditions
were similar for all tested three genotypes of pomegranate. The results were in agreement with various
records in many plants species such as sunflower (Venkatraju and Marschner, 1981), chrysanthemum
(Rutland and Bukovac, 1971), soybean (Fleming et al., 1984), orange (Romheld et al., 1982), apple (Sha-
habi et al., 2005), and cucumber (Romera and Alcantara, 1994). However, Paz et al. (2012) in a study on
Lotus tenuis treated with NaHCO3 recorded that NaC levels and the NaC/KC ratio remained relatively
unchanged, and Zhang et al. (2012) reported that higher NaC exclusion was a non-dependent activity of
plasma membrane HC-ATPase. Disappearance of the role of plasma membrane HC-ATPase may also
lead to the loss of potassium. The decline of this element in roots could act as a signal for the execution of
a cell death program (Huh et al., 2002; Demidchik et al., 2010).
The lowest NaC/KC ratio in leaves and stems was observed in GD cultivar and the highest ratio was
observed in GHA cultivar. It seemed quick absorption of NaC synchronized with decrease of KC con-
centration led to increased NaC/KC ratio in leaves and stems, especially in GHA cultivar. This could
cause disorder in osmosis regulations and could lead to osmosis stress (Bie et al., 2004). Considering
that the lowest root NaC/KC ratio was seen in GD cultivar, it seemed that this cultivar started to
decrease or prevent importing of NaC to leaf from root. Therefore, GD cultivar profited by avoiding
the entry of NaC to root and/or saving imported NaC in root. Consequently, this action prevented NaC
translocation toward shoot. Increase in NaC/KC ratio in leaves of GHA cultivar was because of sensitiv-
ity of this cultivar to the presence of NaHCO3 that led to disorder in exchange activity of the NaC/HC
antiport on the root plasma membrane. In contrast, GD cultivar as a strategy I plant in Fe absorption
had the ability to decrease the effects of NaHCO3 via excretion of phenolic compounds and balancing
the HC import, which is a result of pH balance of the root environment. Increase of NaC in shoots and
roots might be related to decrease in NaC exclusion that has been reported in lettuce by Bie et al.
(2004) and in watermelon grafted onto squash by Colla et al. (2010).

Conclusion
The results of this study showed the reaction of pomegranate to alkalinity was dependent on NaHCO3
critical concentrations in duration of treatment and type of pomegranate cultivar or genotype. Generally,
GD cultivar showed less sensitivity than ZY cultivar to NaHCO3 treatments, if equipped with characteris-
tics such as higher Fe translocation, higher chlorophyll content in upper leaves, and lower NaC levels,
which is a standard for alkalinity tolerance and considering that the used NaHCO3 concentrations did
not have toxic and destructive effects on GD cultivar. However, the response of GD and ZY cultivars to
NaHCO3 in some cases did not show significant difference. Therefore, for presenting GD cultivar as less
 20 H. R. KARIMI AND F. E. TARI

sensitive, ZY cultivar as moderately sensitive, and GHA as sensitive pomegranate cultivars to water irriga-
tion alkalinity, more studies are needed for testing higher concentrations of NaHCO3 on these cultivars.

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