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Analytical Methods

Quality of fresh and stored carrots depending on iodine and nitrogen


fertilization
Sylwester Smolen
a,
, Wodzimierz Sady
a
, Iwona Ledwo
_
zyw-Smolen
b
, Piotr Strzetelski
c
,
Marta Liszka-Skoczylas
d
, Stanisaw Ro
_
zek
c
a
Unit of Plant Nutrition, Institute of Plant Biology and Biotechnology, Faculty of Horticulture, University of Agriculture in Krakw, Al. 29 Listopada 54, 31-425 Krakow, Poland
b
Unit of Biochemistry, Institute of Plant Biology and Biotechnology, Faculty of Horticulture, University of Agriculture in Krakw, Al. 29 Listopada 54, 31-425 Krakow, Poland
c
Unit of Botany and Plant Physiology, Institute of Plant Biology and Biotechnology, Faculty of Horticulture, University of Agriculture in Krakw, Al. 29 Listopada 54, 31-425
Krakow, Poland
d
Department of Engineering and Machinery in Food Industry, Faculty of Food Technology, University of Agriculture in Krakw, ul. Balicka 122, 30-149 Krakow, Poland
a r t i c l e i n f o
Article history:
Received 25 February 2013
Received in revised form 4 November 2013
Accepted 7 March 2014
Available online 15 March 2014
Keywords:
Iodine
Biofortication
Post-harvest physiology
Carrot
Phenolic compounds
DPPH
Carotenoids
a b s t r a c t
Introduction: Iodine is an important mineral nutrient essential for a proper functioning of human and
animal organism. Despite current programmes of iodine prophylaxis (mainly based on salt iodization)
approximately 3038% of human population has insufcient iodine intake. Crop plants can become an
efcient vector of this element in the food chain. Iodine is not a nutrient for plants. For that reason, in
addition to determining the possibility of increasing iodine content in crop plant it is necessary to
describe its impact on yield quality. The aim of the study was to analyze the inuence of soil fertilization
with iodine and nitrogen on the quality of carrot roots and its storage ability.
Methods: In 20082010 the eld study with carrot cv. Kazan F
1
was conducted. A differential soil fertil-
ization with iodine (in the form of I

or IO
3

) and nitrogen (as NO


3

or NH
4
+
) was applied in the experiment:
(1) control without N and I, (2) KI application without N, (3) KIO
3
application without N, (4) KI + Ca(NO
3
)
2
,
(5) KIO
3
+ Ca(NO
3
)
2
, (6) KI + (NH
4
)
2
SO
4
and (7) KIO
3
+ (NH
4
)
2
SO
4
. The experiment was arranged in a split-
plot design. Iodine (in both forms) was applied pre-sowing in a dose of 2 kg I ha
1
. Nitrogen in the form of
Ca(NO
3
)
2
and (NH
4
)
2
SO
4
was introduced pre-sowing and as a top dressing, each dose of 100 kg N ha
1
.
Results and discussion: A diverse, statistically signicant inuence of tested factors on the activity of free
radical-scavenging (DPPH) and the content of: dry matter, glucose, fructose, sucrose, total soluble sugars,
soluble solids Brix %, phenolic compounds, phenylpropanoids, avonols, anthocyanins and carotenoids
was noted in carrot roots directly after the harvest as well as at the end of four-month storage. Iodine
applied with relatively high doses of nitrogen decreased the quality of fresh carrot. After storage, opposite
relations were noted for tested combinations (with I and N application) with respect to carrot quality
when compared to results obtained after the harvest. The lowest storage ability was found for carrot trea-
ted with KI without N. Obtained results directly suggest the need for developing individual agronomic
rules for iodine biofortication of carrot for: (a) consumption and/or processing directly after the harvest
and (b) long-term storage.
2014 Elsevier Ltd. All rights reserved.
1. Introduction
Iodine is an important mineral nutrient necessary for the proper
functioning of human body. It is involved in the biosynthesis of
thyroid hormones [thyroxine (T4) and triiodothyronine (T3)] that
regulate numerous metabolic processes in the whole organism.
In medicine, the spectrum of developmental and functional disor-
ders caused by too low an intake of iodine are dened as iodine
deciency disorders (IDD). In the pre- and neonatal period, this
nutrient has a crucial role in neurological and brain development
(Melse-Boonstra & Jaiswal, 2010; Walker et al., 2007).
It is estimated that approximately 3038% of human population
has inadequate iodine intake and is, therefore, at risk of IDD (White
& Broadley, 2009; Winger et al., 2008). Europeans have notably
poor iodine intake with 56.9% children and ca. 60% adults consum-
ing less than the recommended daily intake (Andersson, de
Benoist, Darnton-Hill, & Delange, 2007; de Benoist, Andersson, Egli,
Takkouche, & Allen, 2004) despite numerous public health
programmes, based mainly on salt iodization, in most member
http://dx.doi.org/10.1016/j.foodchem.2014.03.024
0308-8146/ 2014 Elsevier Ltd. All rights reserved.

Corresponding author. Tel.: +48 12 6625239; fax: +48 12 6625240.


E-mail address: s.smolen@ogr.ur.krakow.pl (S. Smolen ).
Food Chemistry 159 (2014) 316322
Contents lists available at ScienceDirect
Food Chemistry
j our nal homepage: www. el sevi er . com/ l ocat e/ f oodchem
states of the European Union. Low intakes result from poor iodine
stability in salt and losses during production, packaging, transpor-
tation and processing. The total amount of iodine lost from salt
may be up to 90% with cooking alone contributing to approxi-
mately 20% of these losses (Winger et al., 2008).
Plant biofortication is proposed as a fast and relatively low-
cost method of introducing iodine into the food chain (White &
Broadley, 2009; Yang, Chen, & Feng, 2007). Vegetables with
increased iodine content could be the preferred vector for this min-
eral in the diet as many of these food products are consumed raw
(Haldimann, Alt, Blanc, & Blondeau, 2005). Iodine is not considered
to be a plant nutrient (Kabata-Pendias, 2011), which is a signicant
factor limiting application in plant fertilization strategies.
Numerous studies have been conducted so far on iodine uptake
and accumulation in plants (Mackowiak & Grossl, 1999; Weng,
Hong et al., 2008; Weng, Weng et al., 2008; Ujowundu et al.,
2011). An interesting approach is the utilization of biotechnology
methods for increasing iodine content in plants through the inhibi-
tion of its methylation and volatilization of methyl iodide /CH
3
I/ to
the atmosphere (Landini et al., 2012).
Previous studies on iodine biofortication of carrot have been
conducted in greenhouse conditions with the application of rela-
tively high iodine doses (Dai, Zhu, Zhang, & Huang, 2004; Hong,
Weng, Qin, Yan, & Xie, 2008). No results are available documenting
the effect of fertilization with iodine on carrots grown in elds
using standard agricultural practices. What is more, no studies
have been conducted so far on plant biofortication with iodine
and its effects on post-harvest physiology of crop plants. Thus, it
seems necessary not only to compare the efciency of various
methods for increasing iodine content in plants, but also to discuss
its inuence on crop quality.
The aim of this study was to determine the effect of different
iodine (potassium iodide or potassium iodate) fertilizers in parallel
with nitrogen (calcium nitrate or ammonium sulphate) on carrot
quality and storage.
2. Materials and methods
2.1. Plant material and treatments
During 20082010, a eld study with carrot cv. Kazan F
1
was
conducted in the Experimental Station (50
o
07
0
910 N, 19
o
84
0
764
E) of University of Agriculture in Krakow Poland, each year on a
different site within a single soil complex.
Different soil fertilization with iodine (in the form of I

or IO
3

)
and nitrogen (as NO
3

or NH
4
+
) was applied in the experiment: (1)
control without N and I, (2) KI application without N, (3) KIO
3
application without N, (4) KI + Ca(NO
3
)
2
, (5) KIO
3
+ Ca(NO
3
)
2
, (6)
KI + (NH
4
)
2
SO
4
, (7) KIO
3
+ (NH
4
)
2
SO
4
. The experiment was ar-
ranged in a split-plot design. Each treatment was randomised in
four repetitions on 2.7 5 m (13.5 m
2
) plots. The total area of
the experiment was 378 m
2
.
Iodine (in both forms) was applied pre-sowing in a dose of
2 kg I ha
1
as pure salts (KI POCH Poland, KIO
3
Sigma Aldrich

).
Nitrogen in the form of Ca(NO
3
)
2
[Yara International ASA (Hydro)]
and (NH
4
)
2
SO
4
[Zakady Azotowe in Tarnw, Poland] was intro-
duced in two 100 kg N ha
1
doses: pre-sowing and as a top-dress-
ing. Pre-sowing application of nitrogen and iodine was conducted
before ridge formation, whereas the second dose of N at canopy
closure (27 June 08, 26 June 09 and 07 July 10). Carrots were cul-
tivated in one row on 40 cm wide and 30 cm high ridges at a seed-
ing rate of 37 seeds m
1
(approximately 550 000 seeds per
hectare). The seeds were sown on 24 April 08, 24 April 09 and 23
April 10. The carrot roots were harvested on 30 September 08, 23
September 09 and 30 September 10. At harvest, approximately
5 kg samples of carrot storage roots were chosen from each of
the four plots (replications) for laboratory analysis. Additionally,
for each of the experimental treatments, a mixed sample of 25 kg
of roots was collected for storage. During harvest, mean carrot
length was also determined. Only roots qualifying as marketable
yield were taken for further analyses. Marketable yield consisted
of storage roots of cylindrical or close-to-cylindrical shape with
head diameter of P3 cm, undamaged by pests, not infected by fun-
gi or bacteria, with no fractures and heads greened to a maximum
of 0.5 cm. The length of a storage root was 15 cm minimum.
2.2. Storage conditions
Stored carrots from each year of the study were placed in plastic
boxes (60 40 25 cm) and kept at normal atmosphere, 1 C, and
9598% humidity. After four-months, healthy roots (not affected
by diseases) were weighed and taken for analysis.
2.3. Plant analysis after harvest and long-term storage
Stored roots were washed and juiced or diced using a household
processor immediately before analysis. The content of total soluble
solids (Brix) was measured using a Atago Palette PR-32 a digital
refractometer. Dry matter was assessed at 105 C. Levels of glu-
cose, fructose, sucrose and total sugars (calculated) were deter-
mined using RP-HPLC. Determination of these compounds was
conducted in room temperature using a Knauer system (Germany).
Samples (10 ll) were injected on an amine LiChrospher RP 100-10
NH
2
250 4 mm column. The eluent was a mixture of acetonitryl/
water (87:13 v/v) and the ow rate 1.3 ml/min. Measurement was
conducted using Smartline RI refractometric detector 2300 (Bogdanov,
2002). Total carotenoid content was assayed after sample extrac-
tion with acetone/n-hexane (4:6) using a b-carotene standard
curve.
To estimate phenolic constituents and free radical scavenging,
carrot extracts were prepared in 80% methanol. Total phenols, phe-
nylpropanoids, avonols and anthocyanins concentrations were
determined using the spectrophotometric method described by
Fakumoto and Mazza (2000). Free radical scavenging activity was
evaluated on the basis of 30-min plant tissue reaction with diph-
enylpicrylhydrazyl (DPPH) (Pekkarinen, Stockmann, Schwarz,
Heinnonen, & Hopia, 1999).
2.4. Statistical analysis
Results were statistically veried using ANOVA module of Stat-
istica 9.0 PL programme for signicance level P < 0.05. Signicance
of changes was assessed with the use of variance analysis F test.
In case of signicant changes, homogenous groups were distin-
guished on the basis of Duncan test.
All data were presented as means for the years 20082010 as in
each year comparable results of treatments on quality of carrot
were obtained.
3. Results and discussion
3.1. Root weight and dry matter content
Results of numerous studies indicate that IO
3

introduced into
the soil or nutrient medium is less harmful for plants than I

when
applied in the same concentrations (Mackowiak & Grossl, 1999;
Blasco, Rios, Cervilla et al., 2011; Blasco, Rios, Leyva et al., 2011;
Caffagni et al., 2011; Hong et al., 2012). In our research no signi-
cant effect of iodine and nitrogen fertilization was found with re-
spect to root weight at harvest and after long-term storage
S. Smolen et al. / Food Chemistry 159 (2014) 316322 317
(Table 1). In our study, the lack of impact on carrot root weight
suggests that soil application of iodine at 2 kg I ha
1
as either I

or IO
3

did not affect the carrot plants negatively. Typically, such


a negative impact would manifest as chlorosis of older leaves,
necrosis and plant death (Hong, Weng, Yan, & Islam, 2009;
Mackowiak & Grossl, 1999; Mackowiak, Grossl, & Cook, 2005).
The strength of this effect depends on iodine dose, chemical form
and method of application, type of plant cultivation (in eld or
protected) and inter-species differences (Caffagni et al., 2011;
Mackowiak et al., 2005; Weng, Hong et al., 2008; Weng, Weng
et al., 2008).
A reduction of carrot root weight was observed after storage
(Table 1). It was mainly related to water evaporation from roots
increasing apparent dry matter content when compared to values
noted at harvest. The loss of weight during long-term storage is a
common observation and the extent depends on agricultural prac-
tices applied during plant cultivation (including dose of nitrogen
fertilization) and varietal traits as well as the length and conditions
of storage (Gajewski, Szymczak, & Danilcenko, 2010; Ro_ zek, Leja, &
Wojciechowska, 2000). The decrease in root weight and changes in
its chemical composition are also related to metabolic processes
occurring during long-term storage, such as cell respiration
(Hajirezaei & Stitt, 1991) or phenolic compound biosynthesis (Leja,
Ro_ zek, & Mareczek, 1998). In the present study the smallest loss of
a total carrot weight as well as the lowest increase of dry matter con-
tent was noted in the control. All treatments with iodine and nitro-
gen fertilization were associated with a loss of carrot root weight
during its storage with the greatest change observed with
KI + Ca(NO
3
)
2
application. Introductionof I

, rather thanIO
3

without
nitrogenelicited the greatest water loss fromcarrots during storage.
This observation was reected in higher dry matter content at its
highest after 4 months. Most likelytheserelationships aretheconse-
quence of processes occurring during carrot cultivation and storage.
3.2. Quality parameters of carrot at harvest
Fertilizer with iodine and nitrogen had a statistically signicant
inuence on quality parameters determined directly after harvest
including the content of: glucose, fructose, sucrose, total sugars,
Brix (Fig. 1AE), phenolic compounds, phenylpropanoids, avo-
nols, anthocyanins (Fig. 2AD), carotenoids (Fig. 3B) as well as free
radical scavenging activity (Fig. 3A). A general trend of increasing
glucose, fructose, total sugars and soluble solids (Brix) together
with a lower content of phenolic compounds and phenylpropa-
noids was noted for KI and KIO
3
application (without N) when
compared to the control. Additionally, it should be underlined that
the highest levels of total sugars and total soluble solids were
noted in carrot roots from these combinations.
At harvest soil fertilization with KIO
3
+ Ca(NO
3
)
2
and KIO
3
+
(NH
4
)
2
SO
4
signicantly decreased levels of: glucose, sucrose, total
sugars, total soluble solids (Fig. 1A, C, D and E) and avonols in
comparison to the control (Fig. 2C). In carrot roots treated with
KIO
3
+ (NH
4
)
2
SO
4
, a signicantly higher content of sucrose, total
soluble solids (Fig. 1C and D) and carotenoids (Fig. 3B) while lower
lever of fructose was noted after application of KIO
3
+ Ca(NO
3
)
2
. In
addition, the content of fructose in carrot grown using this combi-
nation was the lowest in the whole study (Fig. 1B). The lowest con-
centration of phenylpropanoids was observed in carrots treated
with KIO
3
+ (NH
4
)
2
SO
4
while of carotenoids in roots grown with
KIO
3
+ Ca(NO
3
)
2
.
Results of this study indicate the inuence of both forms of io-
dine on carrot quality assessed directly after harvest was depen-
dent on the chemical form of nitrogen including other elements
(Ca
2+
and SO
4
2
), i.e. type of nitrogen fertilizer. Soil fertilization with
ammonium sulphate did not signicantly decrease the content of
sugar compounds in carrot in comparison to the application of cal-
cium nitrate. This is particularly interesting as there is common
agreement that the process of SO
4
2
reduction and its incorporation
into sulphur-containing amino acids require metabolic energy
(Cram, 1983; Leustek & Saito, 1999). In addition, carrot requires
low sulphur, which may explain its low uptake data presented
previously (Smolen , Sady, Ro_ zek, Ledwo_ zyw, & Strzetelski, 2011).
It is striking, however, that application of KIO
3
together with either
Ca(NO
3
)
2
or (NH
4
)
2
SO
4
contributed to a decrease in carbohydrate
content (glucose, fructose, sucrose and total sugars) and total
soluble solids (Fig. 1AE) when compared to KI application with
nitrogen fertilizers. This can be explained based on the theory
IO
3

to I

reduction in plant roots is at the expense of energy re-


leased during cell respiration, prior to transport of iodine to
above-ground parts of plant (Cseh & Bszrmnyi, 1964; Muramatsu,
Christoffers, & Ohmom, 1983). On this basis, it could be hypothe-
sised that nitrate reductase (NR) is involved in IO
3

reduction
(Hung, Wong, & Dunstan, 2005; Wong & Hung, 2001). In the case
of plants fertilised with both forms of nitrogen, the observed ef-
fects could be explained by increased energy consumption by the
reduction of NO
3

(taken from soil) to NH


4
+
and incorporation of
these ions in amino acids and subsequent protein biosynthesis.
These reactions could also decrease total sugar content in carrots
fertilised with iodine and nitrogen compared to plants treated with
KI and KIO
3
without nitrogen. This assumption seems to be reason-
able as, in carrots fertilised with nitrogen, the content of N-total
was substantially higher than in plants cultivated without N
(Smolen et al., 2011). Therefore, more energy may have been con-
sumed in the metabolism of nitrogen and intermediates necessary
for amino acid biosynthesis. It seems results from the present
study go some way to explaining the range of iodine inuence on
sugar metabolism in carrots. In the case of carbohydrates, applica-
tion of IO
3

led to a decrease in glucose, fructose and sucrose in pro-


portion to I

concentration (Blasco, Rios, et al., 2011). Iodine also


appears to stimulate conversion of simple sugars to polysaccha-
Table 1
Root weight and dry matter content in carrot roots at harvest and after storage depending on iodine and nitrogen fertilization (n = 12).
Treatment Average root weight (g) Dry matter content (%)
At harvest After storage Change after storage (%) At harvest (a) After storage (b) Change after storage (ba)
Control 178.1 141.9 20.3 a 11.6 e 13.3 a +1.70
KI without N 175.7 133.3 24.1 d 11.8 f 15.9 b +4.10
KIO
3
without N 186.8 139.5 25.3 f 11.6 e 14.1 a +2.50
KI + Ca(NO
3
)
2
160.7 113.7 29.2 g 11.2 c 13.9 a +2.70
KIO
3
+ Ca(NO
3
)
2
153.9 121.3 21.2 b 10.6 a 13.0 a +2.40
KI + (NH
4
)
2
SO
4
152.3 118.8 22.0 c 11.3 d 14.3 a +3.00
KIO
3
+ (NH
4
)
2
SO
4
157.0 118.9 24.3 e 11.1 b 13.3 a +2.20
Test F for treatments n.s. n.s.


Test F:

means are signicantly different; n.s. differences are not signicant for P < 0.05.
Means followed by the same letters are not signicantly different.
318 S. Smolen et al. / Food Chemistry 159 (2014) 316322
rides in barley plants that manifested as the accumulation of poly-
saccharides in plant stems, which contributed to higher mechani-
cal stability (Szkolnik, 1980).
To obtain a better picture of the impact of iodine and nitrogen
on carrot quality, a pot experiment was conducted in 2008 using
the same cultivar, soil type and experimental design as presented
in this work. Results describing the chemical analysis of carrots
at harvest are presented elsewhere (Smolen , Ledwo_ zyw et al.,
2009; Smolen , Sady et al., 2009). However, in that work, applica-
tion of KI and KIO
3
on plants cultivated without nitrogen fertiliza-
tion caused a signicant decrease in the content of soluble solids
(Brix), carotenoids (Smolen, Ledwozyw, Strzetelski, Sady, and
Rozek, 2009) and total phenolic compounds as well as phenylprop-
anoids, avonols and anthocyanins (Smolen, Sady, Strzetelski,
Rozek, and Ledwozyw, 2009). Thus, we obtained similar results
to those of (Smolen, Sady, et al., 2009) with respect to the effect
of iodine (without N) on the accumulation of phenolic compounds
1 2 3 4 5 6 7
0
500
1000
1500
2000
G
l
u
c
o
s
e
(
m
g

1
0
0
g
-
1
f
.
w
.
)
a a
b
bc cd
e f
f
g
h
i
f
d cd
A
1 2 3 4 5 6 7
0
500
1000
1500
2000
F
r
u
c
t
o
s
e
(
m
g

1
0
0
g
-
1
f
.
w
.
)
a b b
c c
d e
f
f
g
h
B
h
c c
1 2 3 4 5 6 7
0
500
1000
1500
2000
2500
3000
3500
S
u
c
r
o
s
e

(
m
g

1
0
0
g
-
1
f
.
w
.
)
C
a
a
b
c
d e
f
f
g g
g
b
b
b
1 2 3 4 5 6 7
0
1000
2000
3000
4000
5000
6000
7000
T
o
t
a
l
s
u
g
(
m
g

1
0
0
g
-
1
f
.
w
.
)
D
a
b
c
d
e
f
g h i
j k
l
de
a
a
r
s
1 2 3 4 5 6 7
0
2
4
6
8
10
12
14
16
B
r
i
x
(
%
)
E
a
b
c c d
e
f
g h h i
j k
l
At harvest After storage
At harvest After storage
Fig. 1. Content of: glucose (A), fructose (B), sucrose (C), total sugars (D) and total
soluble solids (E) in carrot roots at harvest and after storage depending on iodine
and nitrogen fertilization. Treatments: 1 control without N and I fertilization;
2 KI fertilizationwithout Napplication; 3 KIO
3
fertilization without Napplication;
4 KI+Ca(NO
3
)
2
fertilization; 5 KIO
3
+Ca(NO
3
)
2
fertilization, 6 KI+(NH
4
)
2
SO
4
fertilization, 7 KIO
3
+(NH
4
)
2
SO
4
fertilization. Means followed by the same letters are
not signicantly different for P < 0.05 (n = 12).
1 2 3 4 5 6 7
0
10
20
30
40
50
60
70
80
P
h
e
n
o
l
i
c
c
o
m
p
o
u
n
d
s
(
m
g

1
0
0
g
-
1
f
.
w
.
)
A
a
ab
bc bcd
d
e
ef
fg
gh
hi
i
j
cd
ab
1 2 3 4 5 6 7
0
5
10
15
20
P
h
e
n
y
l
p
r
o
p
a
n
o
i
d
s
B
a ab ab
abc bc
bc
d
d
e
g
h
f
e
c
(
m
g

1
0
0
g
-
1
f
.
w
.
)
1 2 3 4 5 6 7
0
2
4
6
8
10
12
F
l
a
v
o
n
o
l
s
C
a
b
c
d
e
d
c c
c
b
b
ab a a
(
m
g

1
0
0
g
-
1
f
.
w
.
)
1 2 3 4 5 6 7
0.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
A
n
t
h
o
c
y
a
n
i
n
s
D
a
a
a a
a
a
a
a
ab
b
ab
b
b
b
(
m
g

1
0
0
g
-
1
f
.
w
.
)
At harvest After storage
At harvest After storage
Fig. 2. Content of phenolic compounds (A), phenylpropanoids (B), avonols (C) and
anthocyanins (D) in carrot roots at harvest and after storage depending on iodine
and nitrogen fertilization. Treatments: 1 control without N and I fertilization;
2 KI fertilization without Napplication; 3 KIO
3
fertilizationwithout Napplication;
4 KI+Ca(NO
3
)
2
fertilization; 5 KIO
3
+Ca(NO
3
)
2
fertilization, 6 KI+(NH
4
)
2
SO
4
fertilization, 7 KIO
3
+(NH
4
)
2
SO
4
fertilization. Means followed by the same letters are
not signicantly different for P < 0.05 (n = 12).
S. Smolen et al. / Food Chemistry 159 (2014) 316322 319
in carrot (Fig. 2AD). Particularly of interest is the fact that, in the
pot experiment, no signicant inuence on the content of soluble
sugars was found (Smolen, Ledwozyw, et al., 2009). Additionally,
treatment with KIO
3
together with calcium nitrate and ammonium
sulphate led to a signicant decrease [for Ca(NO
3
)
2
] or increase [for
(NH
4
)
2
SO
4
] in the content of soluble solids, phenolic compounds,
phenylpropanoids and avonols in carrot when compared to KI
application with the same nitrogen fertilizers (Smolen, Ledwozyw,
et al., 2009; Smolen, Sady, et al., 2009). In the case of total sugars
and soluble solids, results obtained by (Smolen, Ledwozyw, et al.,
2009) were different to those presented here (Fig. 1D, E). This dis-
crepancy may be due to different growing conditions, e.g. eld ver-
sus pot cultivation. In reference to the impact of I and N on levels of
total phenolic compounds, phenylpropanoids, and avonols, it is
noteworthy that iodine as I

and IO
3

did not stress the plants. In-


creased biosynthesis of these compounds in plants may be ob-
served as abiotic stress (Korkina, 2007; Michalak, 2006) and may
be triggered by excessive iodine (Blasco et al., 2008). The efciency
of iodine biofortication was 588.6740.1 lg I kg
1
d.w for
KI + (NH
4
)
2
SO
4
and KIO
3
+ Ca(NO
3
), respectively, while the content
of iodine in control plants was 240.1 lg I kg
1
d.w. Detailed results
including biofortication level, yielding, iodine transport from
roots to leaves and changes in iodine root content during long-
term storage were presented in the dissertation by Smolen (2013).
3.3. Effect of storage on carrot quality
Our results suggest iodine biofortication of carrot can substan-
tially affect post-harvest physiological and biochemical processes
in storage roots. A statistically signicant inuence of factors
affecting quality parameters in carrot was noted after long-term
storage (Fig. 1AE, Fig. 2AD, Fig. 3A and B). Carrots grown under
all combinations (including the control) had a signicantly higher
level of scavenging activity (Fig. 3A), glucose, fructose, total sugars,
total soluble solids (Fig. 1A, B, D and E), phenolic compounds,
phenylpropanoids, avonols (Fig. 2A, B and C) and carotenoids
(Fig. 3B) while lower levels of sucrose (Fig. 1C) were found after
long-term storage than immediately after harvest. It is most likely
this effect is due to water evaporation (direct loss of carrot weight).
As a consequence, quantitative relationships between fertilizer
combinations and quality parameters (after storage) were substan-
tially different from those found after harvest. It seems the cause of
these changes cannot be solely ascribed to water loss (expressed as
differences in dry matter content before and after storage), but also
to a specic effects of iodine and nitrogen on post-harvest physio-
logical and biochemical processes in carrot.
In comparison to the control, a signicant decrease in the accu-
mulation of fructose following KI + (NH
4
)
2
SO
4
application and total
soluble solids with KIO
3
+ (NH
4
)
2
SO
4
as well as sucrose content in
roots fertilised with KIO
3
+ Ca(NO
3
)
2
compared to controls. Carrots
grown under all combinations (except for KI application without
N) contained signicantly less phenolic compounds, phenylpropa-
noids and avonols, and were characterised by improved scaveng-
ing activity compared to controls.
In higher plants, iodine can occur as iodised amino acids, mainly
derivatives of aromatic amino acids such as tyrosine and thyronine
(Szkolnik, 1980). In addition, it has been noted that most iodine
accumulated in marine and terrestrial plants is bound to proteins
(Burianov, Macht, Niedobov, Doucha, & Kanicky , 2005; Hou,
Yan, & Chai, 2000; Weng et al., 2008). Details of its structure and
role in plant organisms have not been elucidated. In our study par-
ticularly interesting was the signicant increase in the levels of
phenolic compounds, phenylpropanoids and avonols in roots
grown with KI but not nitrogen. This could have been related to
the unfavorable effect of I

on physiological processes in root cells


of plants with lower nitrogen status and, therefore, decreased pro-
tein synthesis. It seems this combination may trigger post-harvest
stress, leading directly to enhanced synthesis of phenolic com-
pounds, phenylpropanoids and avonols (Fig. 2AC), greater loss
of root weight and increased dry matter content (Table 1). To some
extent this suggestion may be supported by the fact that, after stor-
age, lower levels of phenolic compounds, phenylpropanoids and
avonols as well as glucose, fructose, sucrose, total sugars and total
soluble solids were noted in carrots treated with KIO
3
[with or
without N fertilizer Ca(NO
3
)
2
]. Our results indicate different pro-
cesses may be involved in sugar metabolism during storage in
plants fertilised with KI and KIO
3
. It is possible that IO
3

application
during carrot cultivation stimulates catabolic processes that use
carbohydrates substrates during post-harvest storage. There is a
signicant linkage between these processes and nitrogen fertiliza-
tion of plants. Currently, it is not possible to suggest a reason for
this, as knowledge of iodine inuence on plant metabolism re-
mains insufcient. It is not unlikely that the introduction of iodine
in different oxidation states and, therefore, characterised by vari-
ous levels of toxicity triggers adverse physiological and biochemi-
cal response in plants. The process of iodine methylation followed
by the synthesis and volatilization of CH
3
I may be involved in plant
detoxication fromexcessive iodine (Landini et al., 2012; Muramatsu
& Yoshida, 1995). Generally, the activity of S-adenosylmethionine-
dependent halide/thiol methyltransferase (HTMT) an enzyme
responsible for iodine methylation is greater in leaves than roots
(Itoh et al., 2009). However, there is no direct evidence suggesting
that CH
3
I synthesis by HTMT can occur during storage. Enzymes
extracted from suspension culture catalysed methylation of caf-
feoyl-CoA to feruloyl-CoA in the presence of S-adenosyl-L-methio-
nine, which determines the activity of HTMT. This process was
described as related to plant defense against Phytophthora mega-
sperma f. sp. Glycinea infection (Khnl, Koch, Heller, & Wellmann,
1989). On the basis of this information, it can be supposed that
during long-term storage carrot carbohydrates are used not only
in catabolic processes (mainly cell respiration), but also in the syn-
1 2 3 4 5 6 7
0
2
4
6
8
10
12
D
P
P
H
(
%
)
A
a
ab
b
c
c
d
e
f
e
de
b
b a b a ab
1 2 3 4 5 6 7
0
5
10
15
20
25
30
C
a
r
o
t
e
n
o
i
(
m
g

1
0
0
g
-
1
f
.
w
.
)
B
a
ab bc bc
bc c
d de de
e
f
f
c
bc
d
s


At harvest After storage
At harvest After storage
Fig. 3. Free radical scavenging activity (A) and content of carotenoids (B) in carrot
roots at harvest and after storage depending on iodine and nitrogen fertilization.
Treatments: 1 control without N and I fertilization; 2 KI fertilization without N
application; 3 KIO
3
fertilization without N application; 4 KI+Ca(NO
3
)
2
fertilization; 5 KIO
3
+Ca(NO
3
)
2
fertilization, 6 KI+(NH
4
)
2
SO
4
fertilization,
7 KIO
3
+(NH
4
)
2
SO
4
fertilization. Means followed by the same letters are not
signicantly different for P < 0.05 (n = 12).
320 S. Smolen et al. / Food Chemistry 159 (2014) 316322
thesis of various compounds, including methyl iodide and second-
ary metabolites important for plant defense against storage
diseases.
4. Conclusion
Based on the results from our study with Kazan F
1
cultivar, it is
possible to assess to some extent the efciency of iodine biofortif-
ication of carrot taking into consideration its inuence on yield
quality and storage potential of this crop. Increasing iodine content
in carrot may be advisable due to a high level of b-carotene a di-
rect precursor of vitamin A that has been shown to have a positive
effect on thyroid function (Zimmermann, 2007). Iodine application
combined with plant fertilizer with relatively high doses of nitro-
gen (100 kg N ha
1
applied pre-sowing and as a top dressing in
the form of calcium nitrate and ammonium sulphate) did, how-
ever, decrease the quality of carrot yield evaluated at harvest. As
far as storage potential is concerned, the lowest value was noted
for carrots grown with KI without N. Analysis of our results sug-
gests there is a need to develop independent methods of iodine
biofortication for carrots intended for (a) consumption and/or
processing directly after harvest or (b) long-term storage. The com-
bination of iodine biofortication programmes with appropriate
crop fertilization with mineral nutrients should also be taken into
account. Furthermore, it seems reasonable to employ plant-breed-
ing techniques to select cultivars characterised by increased iodine
accumulation without decreased yield quality and storage
potential.
Basing on conducted research it can be proposed that for the
cultivation of carrot biofortied with iodine, KI form of this ele-
ment should be applied rather than KIO
3
(also when combined
with nitrogen fertilization) due to a signicantly more deleterious
effect of iodates on carrot quality after harvest. In the production of
carrot for long-term storage of roots high doses of nitrogen fertiliz-
ers need to be avoided (particularly in the form of calcium nitrate)
when applied together with iodine in order to prevent increased
losses of root weight or the decrease in its quality.
Acknowledgement
This work was nanced by research grants from the Ministry of
Science and Higher Education Republic of Poland.
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