Chemical and sensory quality of fresh pomegranate fruits exposed

to gamma radiation as quarantine treatment

Haz Muhammad Shahbaz
a
, Jae-Jun Ahn
a
, Kashif Akram
a,b
, Hyo-Young Kim
a
, Eun-Joo Park
a
,
Joong-Ho Kwon
a,
a
School of Food Science & Biotechnology, Kyungpook National University, Daegu 702-701, Republic of Korea
b
Institute of Food Science and Nutrition, University of Sargodha, Sargodha 40100, Pakistan
a r t i c l e i n f o
Article history:
Received 22 May 2013
Received in revised form 9 August 2013
Accepted 14 August 2013
Available online 26 August 2013
Keywords:
Pomegranates
Gamma-irradiation
Quarantine disinfestation treatment
Chemical properties
Antioxidative capacity
Sensory prole
a b s t r a c t
The U.S. Department of Agriculture in February 2012 approved the import of fresh pomegranates sub-
jected to irradiation as a quarantine procedure with a minimum absorbed dose of 0.4 kGy against differ-
ent pests. This study evaluated the application of different gamma-irradiation doses (0.4, 1, and 2 kGy) in
fresh pomegranate fruits and their effect on the chemical and sensory characteristics. The total soluble
solids, titratable acidity, and pH values remained unaffected up to 1 kGy treatment. Irradiation caused
a signicant decrease in the total anthocyanins and phenolic content. A strong positive correlation was
observed among the antioxidant activities, total phenolics and anthocyanin contents. In general, a stron-
ger preference was shown by sensory panelists for the juice from irradiated fruits. This study provides
research-based information about the application of irradiation as a quarantine disinfestation treatment
to enhance the marketing and consumer acceptance of pomegranates.
2013 Elsevier Ltd. All rights reserved.
1. Introduction
Pomegranates (Punica granatum L.) are one of the important
commercial fruits extensively cultivated in many tropical and sub-
tropical regions of the world (Tehranifar, Zarei, Nemati, Esfandiy-
ari, & Vazifeshenas, 2010). The fruit has wide consumer
preference as the consumption of fresh arils or juice because of
its exceptional and unique sensory and nutritional properties
(Varela-Santos et al., 2012). The arils contain a considerable
amount of polyphenols, polysaccharides, sugars, acids, vitamins,
and important minerals (Al-Maiman & Ahmad, 2002). A growing
number of scientic studies have highlighted the potential human
health related benets of pomegranate juice such as antiathero-
genic, antioxidant, antihypertensive, etc. (Mena et al., 2011;
Rajasekar, Akoh, Martino, & MacLean, 2012; Tehranifar et al.,
2010; Varela-Santos et al., 2012; Zaouay, Mena, Garcia-Viguera, &
Mars, 2012).
The fresh consumption of pomegranates has increased in Korea
but local production, that covers an area of about 161.4 ha, is not
enough to full market demand (Shahbaz, Akram, Ahn, & Kwon,
2013). In the past few years, the United States (U.S.) has captured
the major market share (97%) of all the imported pomegranates in
the Korean market. The fresh pomegranates imported from the U.S.
are preferred by consumers because of their uniformity and
consistency in quality. The Korea Food and Drug Administration
(KFDA) has been authorised to conduct inspections of pomegranate
fruits upon arrival at ports in Korea. According to the KFDA estab-
lished standards and phytosanitary import requirements, the im-
ported fruits must contain proper labelling indicating sufcient
information. In addition, pomegranates should not undergo any
disinfestation treatment such as fumigation, etc. by importers to
qualify for organic certication as Korean consumers perceive
low-chemical products as healthy products (U.S. Agricultural Trade
Ofce, 2010).
Pomegranate fruits have a high risk of infestation with sucking
insects and mite pests during growth which deteriorates their
quality and constrain the international trade (Ananda, Kotikal, &
Balikai, 2009). According to pest risk assessment prepared by the
Animal and Plant Health Inspection Service (APHIS) of the U.S.
Department of Agriculture (USDA, 2012), 25 quarantine pests
including two mites (Tenuipalpus granati and Tenuipalpus punicae)
could follow the pomegranate fruit pathway. The APHIS in Febru-
ary 2012 authorised the import of fresh pomegranates into the
U.S. mainland from India to a minimum irradiation dose of
0.4 kGy as quarantine disinfection treatment. The recommended
irradiation dose, along with standard postharvest processes, will
help to effectively neutralise the concerned insect pests and miti-
gate the risks of their dissemination (USDA, 2012). The most feasi-
ble application of irradiation technology in agricultural products,
including fruits, is probably quarantine disinfestation without sig-
nicantly affecting the chemical or sensory attributes (Fields &
0308-8146/$ - see front matter 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.foodchem.2013.08.052

Corresponding author. Tel.: +82 53 950 5775; fax: +82 53 950 6772.
E-mail address: jhkwon@knu.ac.kr (J.-H. Kwon).
Food Chemistry 145 (2014) 312318
Contents lists available at ScienceDirect
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j our nal homepage: www. el sevi er . com/ l ocat e/ f oodchem
White, 2002). However, research is required to determine the
appropriate irradiation doses and their subsequent effects on dif-
ferent quality attributes in fruits. In general, the FDA restricts the
maximum irradiation dose level to 1 kGy for disinfestation and de-
layed maturation in fresh fruits (Boylston, Reitmeier, Moy, Mosher,
& Taladriz, 2002).
The aim of this study was to investigate the effect of gamma-
irradiation, as a quarantine disinfestation treatment, on the chem-
ical and sensory qualities of fresh pomegranate fruits. In addition,
total polyphenols and antioxidant activities were assessed using
two different radical scavenging assays. The results were used to
test for correlations between different quality-related parameters.
2. Material and methods
2.1. Pomegranate fruits, irradiation, and juice extraction
This study was done using pomegranate fruits grown in Califor-
nia (California cultivar; U.S. origin) and freshly imported to Korea
(December 2012). Seventy commercially available fresh pome-
granates, packed in cardboard boxes, were purchased from a local
market in Daegu, South Korea. The fruits were divided into four
equal portions and labelled with the specic radiation dose.
Approximately, 16 fruits were sampled for each irradiation treat-
ment and kept overnight at 5 C in the laboratory. The packed fruit
samples were then irradiated at the Korea Atomic Energy Research
Institute (Jeongeup, Korea) with doses of 0, 0.4, 1, and 2 kGy using
a Cobalt-60 gamma-ray source (AECL, IR-79, MDS Nordion Interna-
tional Co. Ltd., Ottawa, Ontario, Canada).
The irradiation process was accomplished at room temperature
with a dose rate of 1.5 kGy/hr. The absorbed doses (5.6%) were
calibrated by alanine dosimeters with a 5 mm diameter (Bruker
Instruments, Rheinstetten, Germany) in which the free-radical sig-
nals were determined with a Bruker EMS 104 EPR analyzer (Bruker
Instruments, Rheinstetten, Germany) (Shahbaz et al., 2013). After
irradiation processing, the pomegranate fruits were taken to the
laboratory and manually cut with a sterile sharp blade to separate
the eshy arils. Juice was extracted from the isolated arils using a
solid fruit juice extractor (Juice Extractor, Model Le Duo, Magimix,
France). The extracted juice was pour into labelled sterile glass bot-
tles and immediately analysed or stored at 4 C. The experimental
juice samples were ltered with Whatman qualitative lter paper,
Grade 4, before the chemical analyses. The pure juice samples were
diluted at different proportions with distilled water for different
analyses until the absorbance was within the linear range of the
spectrophotometer (Optizen 2120UV, Mecasys Co. Ltd., Daejeon,
Korea). All the analyses were independently repeated three times
to ensure accuracy. All the chemicals used were of analytical grade
and purchased from SigmaAldrich.
2.2. Chemical analyses
The titratable acidity (TA) of the juice was measured by titrating
it against 0.1 N NaOH to the end point of pH 8.1, monitored with a
pH meter. The results were expressed as percentage of citric acid.
The pH measurements were performed using a digital pH meter
(Orion 3 star, Thermo Electron Co., Waltham, MA, USA) at 21 C.
The total soluble solids (TSS) in the juice were determined with a
digital refractometer (Master-M, ATAGO, Brix 032%, Tokyo, Japan)
at 20 C. The instrument was calibrated with distilled water before
the analysis. TSS values were expressed as Brix (Rajasekar et al.,
2012).
The total anthocyanins content in juice samples was deter-
mined with the pH differential method using two buffer systems:
potassium chloride buffer, pH 1.0 (0.025 M) and sodium acetate
buffer, pH 4.5 (0.4 M) according to Giusti and Wrolstad (2001).
Briey, 1 mL of diluted juice was mixed with 4 mL of correspond-
ing buffers. Absorbance was measured at two wavelengths,
510 nm and 700 nm, after 15 min of incubation in a spectropho-
tometer against distilled water as a blank. The total anthocyanins
content was calculated by applying the standard formula. The val-
ues, three replications per sample, were expressed as mg cyanidin-
3-glucoside per 100 mL of juice.
The juice colour measurements were done in a colorimeter
(CM-3600d, Konica Minolta, Osaka, Japan) using the Hunter Lab
scale (L

: Lightness; a

: redness; b

: yellowness) previously de-

scribed by Rajasekar et al. (2012). The instrument was calibrated
against a white reference plate provided with the chromameter be-
tween different readings. The quartz cell was lled with ltered
juice and colour data were recorded with the Minolta Software
Chroma control data system. The average values of 3 measure-
ments were reported.
The amount of total sugars in the fruit juice was determined
using a modied version of the phenolsulphuric acid assay re-
cently described by Nielson (2010). Accurately 1 mL of the diluted
juice sample was mixed with 1 mL of 5% phenol solution and 5 mL
of 96% sulphuric acid (rapidly added) in each tube. The tubes were
vortexed and allowed to stand at room temperature for 20 min.
The concentrated sulphuric acid converts all non-reducing sugars
to reducing sugars, so the method determines the concentration
of the total sugars present in the sample. A blank was prepared
by substituting distilled water for the juice sample. The absorption
of the characteristic yellow-orange colour produced as a result of
the interaction between the sugars and the phenol was measured
at 490 nm using a spectrophotometer. The typical colour of this
reaction is stable for several hours. The concentration of the total
sugars present in each sample was calculated by referring to a
standard sucrose curve.
The content of the reducing sugars was measured with the Nel-
sonSomogyi method (Somogyi, 1952) with minor modications.
The method is widely used for the quantitative determination of
reducing sugars in biological materials. Four types of required solu-
tions were prepared according to standard procedures with high
accuracy. Arsenomolybdate reagent was incubated at 37 C for
24 h prior to use. The diluted juice sample (0.5 mL) was mixed with
the different solutions as previously described. The absorbance of
the blue colour was read at 520 nm with a spectrophotometer.
The amount of reducing sugars present in the fruit juice sample
was calculated from a standard curve graph drawn using a glucose
solution as the standard. The average results for triplicate determi-
nations were expressed as g/100 mL of juice
.
2.3. Total phenolics content and antioxidant capacities
The concentration of the total phenolics was measured by the
FolinCiocalteu reagent method recently described by Rajasekar
et al. (2012). To each 50 lL of diluted juice, 0.5 mL FolinCiocalteu
reagent and 1.5 mL of 7.5% sodium carbonate were added. The
samples were allowed to stand at room temperature for 30 min
incubated under dark conditions. The wavelength of spectropho-
tometer was xed at 765 nm for the absorbance reading. Results
were expressed as mg of Gallic acid/100 mL of juice using a gallic
acid (00.1 mg/mL) standard curve.
Several assays were done to estimate the antioxidant activity in
fresh fruits and their products. Most of the natural antioxidants are
multifunctional; therefore, for a more reliable evaluation, it is
important to perform different antioxidant activity assessments
to give proper consideration to the various mechanisms of antiox-
idant action. In this study, DPPH (2,2-diphenyl-1-picrylhydrazyl)
and ABTS (2,2-azino-bis-3-ethylbenzothiazoline-6-sulphonic acid)
methods were used to measure the antioxidant activity of
H.M. Shahbaz et al. / Food Chemistry 145 (2014) 312318 313
pomegranate fruit juice samples. The free radical-scavenging
activity of the juices was assessed against the stable 1,1-diphe-
nyl-2-picryl hydrazyl radical (DPPH) according to the method of
Mishra, Ojha, and Chaudhury (2012a) with slight modications.
The DPPH and Trolox solutions were freshly prepared in ethanol
and adjusted to a 1.000 0.002 absorbance at 517 nm. Each of
the 1 mL juice samples was shaken vigorously with 5 mL of freshly
prepared ethanolic solution of DPPH (12 mg DPPH/100 mL etha-
nol). The scavenging activity on the DPPH radical of the resulting
solution was measured (525 nm) after 30 s of reaction in the spec-
trophotometer against an ethanol blank. The DPPH scavenging
ability of the extract was calculated from the equation (Mishra
et al., 2012a).
For the ABTS assay, the procedure described by Re et al. (1999)
was followed. Stock solution of ABTS
+
was prepared by reacting a
7 mM aqueous solution of ABTS with 2.4 mM potassium persul-
phate solution and allowed to stand for 16 h at room temperature
in dark conditions. The resulting solution was diluted in ethanol to
an absorbance of 0.700 0.002 units at 734 nm to obtain ABTS
+
working solution. Each of the 200 lL juice samples was blended
with 4 mL of ABTS
+
working solution for a total reaction time of
5 min. The absorbance was taken at a wavelength of 734 nm
against ethanol as a standard using the spectrophotometer. Trolox
(0.041.25 mg/mL) was used as a standard for the calibration curve
(Re et al., 1999).
2.4. Sensory evaluation
Sensory evaluation was performed by randomly choosing a
group consisting of 30 students belonging to the School of Food
Science & Biotechnology, Kyungpook National University. The age
distribution of the panelists was between 22 and 30 years includ-
ing both men and women. The students had basic knowledge on
the concepts and applications of sensory analysis. Appropriate
information about the nature of the present evaluation was also
provided. Irradiated and control juice samples were placed on
top of white paper plates in a randomised order and identied with
3-digit codes. Approximately 30 mL of each juice sample (8 C) was
presented to the judges with 5 min intervals between samples.
Deionised water and unsalted crackers were provided for palate
rinsing in-between the samples. Panelists rated the intensity of
the sensory attributes including color, avor, aroma, sweetness,
sourness, overall taste, and overall acceptances on a ve-point
scale (5 = most like; 4 = good; 3 = fair; 2 = dislike a little; 1 = most
dislike). The scores from 2.55 were considered acceptable (Kim
& Yook, 2009).
2.5. Statistical analysis
Statistical analyses were performed with the Statistical Analysis
System (SAS) (Version 9.2; SAS Inst. Inc., Cary, N.C., U.S.A) using
one-way analysis of variance ANOVA. Duncans multiple range
(DMR) test was applied to calculate the signicant difference be-
tween different irradiation treatments. Results were expressed as
the average standard deviation. Data were also analysed to deter-
mine whether there was any correlation between different fruit
quality attributes using Pearson correlation.
3. Results and discussion
3.1. Effect of irradiation on total anthocyanins content
Anthocyanins are one of the major groups of pigments account-
able for contributing different colors in fruits, including pomegran-
ate juice, and possible health benets such as dietary antioxidants.
During chemical determination analysis, monomeric anthocyanin
pigments undergo a reversible structural transformation as a func-
tion of pH. The pH differential method calculates the total mono-
meric anthocyanin content based on the structural change of the
anthocyanin chromophore between pH 1.0 and pH 4.5 (Tehranifar
et al., 2010).
From the analysis, a slight gradual decrease in the anthocyanin
content of juice from the gamma-irradiated pomegranate fruit was
identied (Table 1). The total anthocyanins content decreased as
the dose increment increased from 0.4 kGy to 2 kGy. However,
the differences were only statistically signicant for the 2 kGy
treatment compared to the control samples. Comparable to these
results, Alighourchi, Barzegar, and Abbasi (2008) directly exposed
extracted pomegranate juice to ionizing radiation (010 kGy) and
observed a signicant reduction in the total and individual antho-
cyanins content at all applied doses, particularly at higher doses
(3.510 kGy). However, from their study results, irradiation treat-
ment of juice with more than 2 kGy was not recommended due
to the detrimental effects on the total anthocyanins content. The
decrease in the anthocyanins content is mainly due to the degrada-
tion of individual anthocyanins. The stability of diglycosides antho-
cyanins to irradiation was higher than monoglycosides at lower
doses (0.5 and 2 kGy) of gamma irradiation (Alighourchi et al.,
2008). Furthermore, the relative stability of an individual anthocy-
anin depends on its matrix, structural features, and the processing
conditions (Alighourchi et al., 2008; Torskangerpoll & Andersen,
2005).
In contrast, Ayed, Yu, and Lacroix (1999) reported an increase in
the anthocyanins content of grape pomace for increasing irradia-
tion doses with an optimum dose at 6 kGy. The stability of antho-
cyanins against irradiation is related to the juice composition. The
increase in the anthocyanins content of grape pomace can be
attributed to the extraction of bound pigments through the degra-
dation of the cell wall. However, most of the available literature on
the applications of irradiation treatment is restricted to solid foods
and there is limited research available on fruit juices (Alighourchi
et al., 2008).
3.2. Effect of irradiation on titratable acidity, pH, and total soluble
solids
Comparison of the effects of irradiation (0, 0.4, 1 and 2 kGy) on
the different physicochemical parameters of the pomegranate juice
samples are presented in Table 1. The titratable acidity in the juice
samples remained unaffected at 0.4 kGy but a signicant decrease
was observed at 1 kGy and 2 kGy treatments. The main organic
acid accountable for the titratable acidity in pomegranate fruit is
citric acid. Similarly, the pH was unchanged up to a 1 kGy dose le-
vel but a higher dose produced a signicant increase in the value.
The total soluble solids concentration of the pomegranates was not
affected at all by the applied irradiation doses.
Conicting results have been reported about the irradiation ef-
fect on pH, TA and TSS parameter in different fruit juices. The pres-
ent results are in close agreement to the ndings of Fan, Niemera,
Mattheis, Zhuang, and Olson (2005) who did not observe any effect
of irradiation at 0.5 and 1.0 kGy on the TA and pH values of sliced
apples, which were initially treated with 7% calcium ascorbate.
Similarly, Miller and McDonal (1996) reported no differences in
the pH values of blueberries when irradiated with gamma-rays
(0.51.0 kGy). In contrast, Moreno, Castell-Perez, Gomes, Da Silva,
and Moreira (2007) found that irradiation up to 3.2 kGy did not af-
fect the pH in blueberry fruits. Yu et al. (1995) found no differences
in the pH values of electron beam irradiated strawberry fruits up to
2.0 kGy.
314 H.M. Shahbaz et al. / Food Chemistry 145 (2014) 312318
3.3. Effect of irradiation on sugars content
Fruit sweetness is an important aspect of fruit quality. Glucose
and fructose are the most predominant sugars present in all fruits
including the pomegranate fruit. Sugars and acids in fruits signi-
cantly inuence the avor, appearance, chemical and sensory char-
acteristics (Al-Maiman & Ahmad, 2002).
Most of the available literature describes the titration based
LaneEynon method for the determination of sugars in fruit juices
(Tehranifar et al., 2010). However, titration methods have several
disadvantages such as the nal results largely depend on precise
reaction times, temperature and reagent concentration. In addi-
tion, the method is susceptible to interference from other types
of molecules that act as reducing agents. In the present investiga-
tion, spectroscopic procedures based on a calorimetric technique
including the phenolsulphuric acid assay and SomogyiNelson
method were used to quantify the amount of total sugars and
reducing sugars, respectively.
Table 1 shows the changes in the sugar content of the fruit sam-
ples upon irradiation. The total sugars content did not change in
the pomegranate fruit at all the applied irradiation doses. However,
compared to the control, slight variations were observed among
different dose levels for the reducing sugars content. The amount
of reducing sugars varied differently at all applied irradiation
doses. Similar ndings have been reported by Mitchell, McLauch-
lan, Isaacs, Williams, and Nottingham (1992) in which the gamma
irradiation had no effect on the sucrose and fructose content of
custard apples at 75 and 300 Gy but a signicant increase was ob-
served in the glucose levels. In the same experiment, no effect was
observed in the fructose and glucose content in lemons at 75 Gy
but an increase was recorded for the sucrose content. Our ndings
are also in agreement with El-Samahy, Youssef, Askar, and Swailam
(2000) in which no effect from the gamma radiation (0.51.5 kGy)
was observed on the total sugars content of mangoes but the
reducing sugars were slightly increased. Research studies have
shown that there is no substantial effect from irradiation on
macronutrients such as proteins and carbohydrates in plant mate-
rials even up to a dose of 10 kGy (Crawford & Ruff, 1996).
3.4. Effect of irradiation on fruit juice color
Anthocyanin pigments are almost exclusively responsible for the
different colors in fruits. The attractive colour (ruby red) of pome-
granate fruit juice has been an important quality attribute which
inuences the consumer behaviour (Zaouay et al., 2012). It may be
signicant to investigate the change in colour because irradiation
can result in the destruction of pigments in fruit and fruit juices.
Juice colour indices showed signicant statistical differences
among the control and irradiated fruit samples. The results (Ta-
ble 2) show that the control fruit juice had a darker colour than
that of the irradiated fruit samples with signicant differences in
the luminosity dimension scale (Lightness; L

= 0 denotes black
and L

= 100 indicates diffuse white). The redness (a

) and yellow-
ness (b

) indices increased directly with the irradiation dose and

represented signicant differences among the samples at different
dose levels. Altogether, the colour difference values (DE) were sig-
nicantly different among the various irradiation doses. The pres-
ent results are consistent with Boylston et al. (2002) who reported
that the colour of 0.75 kGy irradiated rambutan and orange fruits
tended to be more intense than that of the control fruits visually
evaluated by sensory judges. Opposite to these ndings, a study
done by Mitchell et al. (1992) on mangoes showed a reduction in
the a

values after gamma-rays treatment at 75 and 300 Gy. The

change in colour can be ascribed to a decrease in the polyphenol
oxidase activity by irradiation (Mishra et al., 2012b). Subsequently,
the irradiation application produced an important improvement in
the sensory parameters which was also favored by the sensory
judges for the 0.4 kGy and 1 kGy samples.
3.5. Effect of irradiation on total phenolics content
Phenolic compounds are important because they contribute to
the nutritional and sensory quality of fruits. Phenolic compounds
havebeenwidelystudiedinmanyfruits andtheir activityis believed
to be mainly because of the irredox properties, which play a signi-
cant role in adsorbing and neutralizing free radicals (Zaouay et al.,
2012). Some of these compounds, particularly the avonoids, pro-
vide health benets due to their antioxidant capacity. Pomegranate
fruit juice contains a greater amount of phenolic compounds com-
pared to other fruit juices (Tehranifar et al., 2010). FolinCiocalteu
is a simple and widely used method to estimate total phenols based
on the mechanism of the transfer of electrons from phenolic com-
pounds to the FolinCiocalteu reagent in an alkaline medium.
The effect of irradiation on the total phenolic compounds of
juice samples was signicant. Total phenols showed a linear trend
of decrease with the gradual increase in irradiation dose. However,
the irradiation effect was more prominent at higher doses of 1 kGy
and 2 KGy (Fig. 1 A). In addition, a linear relationship was observed
between the total phenolics content and free radical scavenging
activities of pomegranate juice.
Different results have been published for the irradiation effect
on phenolic compounds in foods. The decrease in phenolic con-
tents of pomegranate juice upon irradiation in present study corre-
lates well with the previous ndings of Song et al. (2006) on kale
juice in which a signicant decrease was found in the total phenols
of fruit juice immediately after irradiation (0, 3, and 5 kGy). Ahn
Table 2
Hunters colour values of juice from gamma-irradiated pomegranate fruit (n = 3).
Colour parameter Irradiation dose (kGy)
0 0.4 1 2
L

Lightness 58.05 0.10

d
57.40 0.06
c
57.08 0.04
b
56.47 0.04
a
a

Redness 28.43 0.07

d
28.80 0.02
c
29.81 0.02
b
30.81 0.05
a
b

Yellowness 18.74 0.05

d
18.79 0.08
c
19.39 0.04
b
20.08 0.03
a
DE 0.00 0.00
d
0.51 0.09
c
1.74 0.14
b
3.09 0.05
a
Values with the same superscript letters (ad) in a row are not signicantly dif-
ferent at p < 0.05.
Table 1
Total anthocyanins content (TAC), titratable acidity (TA), pH, reducing sugars (RS), total sugars (TS) and total soluble solids (TSS) of juice from gamma-irradiated pomegranate
fruit (n = 3).
Dose (kGy) Parameters
TAC (mg/100 mL) TA (% citric acid) pH RS (g/100 mL) TS (g/100 mL) TSS (
o
Brix)
0 30.45 0.86
a
0.24 0.012
a
3.45 0.006
b
16.37 240.6
b
18.82 0.076
a
16.3 0.058
a
0.4 29.61 0.19
ab
0.24 0.015
a
3.45 0.006
b
17.04 277.3
a
18.82 0.076
a
16.3 0.058
a
1 28.83 0.26
b
0.23 0.044
b
3.46 0.006
b
16.68 243.9
ab
18.81 0.076
a
16.3 0.058
a
2 28.33 0.39
c
0.22 0.00
c
3.54 0.012
a
16.47 450.6
b
18.82 0.076
a
16.3 0.058
a
Values with the same superscript letters (ac) in a column are not signicantly different at p < 0.05.
H.M. Shahbaz et al. / Food Chemistry 145 (2014) 312318 315
et al. (2005) also reported a signicant reduction in the phenolic
contents of Chinese cabbage at 1 kGy or above. This phenomenon
may occur due to the immediate oxidation of phenolic compounds
thus playing an antioxidant role by reducing the free radicals and
the reactive oxygen species produced by irradiation.
In contrast, Song et al. (2006) found an increase in the total phe-
nols in irradiated (05 kGy) carrot juice immediately after irradia-
tion. El-Samahy et al. (2000) reported that the concentration of the
total phenolic compounds was higher in irradiated mangoes (0.5
1.5 kGy) compared to the control. One possible reason for this
increasing trend can be ascribed to the ability of b-carotene in these
fruits to scavenge radicals (Song et al., 2006).
3.6. Effect of irradiation on antioxidant activities with DPPH and ABTS
assays
The antioxidant activity index of pomegranate juice was
obtained in terms of its radical scavenging potential by using two
(DPPH and ABTS) in vitro assays. These assays have been exten-
sively used for the quantication of antioxidant potential of pheno-
lic samples from different fruits and vegetables. The DPPH assay
relies on the reduction of DPPH which is a stable free radical with
a characteristic absorption and accepts an electron or hydrogen
radical to become a stable diamagnetic molecule. The deep purple
colour of a freshly prepared DPPH solution fades in the presence of
an antioxidant and is converted to a colourless product resulting in
a decrease in absorbance at 517 nm. The characteristic decrease in
absorbance is mainly due to the scavenging of the radical by hydro-
gen donation (Mishra et al., 2012a). Similarly, the ABTS radical cat-
ion is reactive towards most antioxidants particularly the
phenolics. The blue ABTS radical cation is converted to a colorless
form during its interaction with antioxidants leading to a decrease
in absorbance at 734 nm (Re et al., 1999).
As shown in Fig. 1 B, the DPPH values were signicantly lower
in irradiated samples, especially at higher doses. The difference
was statistically signicant for antioxidant activity between the
1 kGy and 2 kGy treated samples. The minimumirradiated samples
(0.4 kGy) represented a slight decrease in DPPH activity compared
to the control. However, the effect of irradiation on DPPH activity
was highly signicant at the maximum irradiation dose (2 kGy).
The ABTS assay prole also showed a corresponding trend to the
total phenolic compounds upon irradiation. ABTS radical scaveng-
ing ability was slightly lowered after irradiation without any sig-
nicant differences. However, there were no differences for the
ABTS values between the 1 kGy and 2 kGy irradiated fruit juices.
The main antioxidant compounds in pomegranate juice are hydro-
lyzable tannins, but anthocyanins and ellagic acid derivatives also
contribute to the total antioxidant capacity of the juice (Gil, Tom-
s-Barbern, Hess-Pierce, Holcroft, & Kader, 2000).
The results are consistent with the studies by Song et al. (2006)
reporting a signicant decrease in the DPPH radical-scavenging
capacity of kale juice upon irradiation (15 kGy gamma-rays) at
day 1. Kim and Yook (2009) found that irradiation doses of 1 and
2 kGy did not signicantly affect the antioxidant potential in kiwi-
fruits throughout a storage period of 3 weeks. In contrast, the DPPH
scavenging ability has dissimilarity to the works of Song et al.
(2006) who evaluated the effect of gamma-irradiation on carrot
juice during a storage period of 3 days at 10 C. The antioxidant
activity of irradiated carrot juice was higher than that of the con-
trol juice but decreased over the storage time.
3.7. Sensory evaluation
Sensory investigations are an important part of food control
because they help to measure and numerically dene the quality
level perceived by judges or consumers. The information about
the sensory quality attributes plays a signicant role in consumer
satisfaction and hence inuences further consumption (Escribano,
Sanchez, & Lazaro, 2010). High doses of irradiation can induce an
off-odour called irradiation odor in fruit juices. Volatile sulphur
compounds, such as hydrogen sulphide, methanethiol, methyl sul-
phide, dimethyl disulphide, and dimethyl trisulphide, play a signif-
icant role in the development of the off-odor. It is observed that
irradiation exerts its effect by hydrolysis of water in foods where
water is a dominant component. Irradiation of water generates
three primary free radicals: hydroxyl, hydrogen atoms, and
hydrated electrons. Use of specic scavengers in a model system
revealed that hydroxyl radicals are involved in the formation of
volatile sulphur compounds (Fan, Lee, & Ahn, 2011). In addition,
0
20
40
60
80
d
c
b
2 1 0.4
T
o
t
a
l

p
h
e
n
o
l
i
c

c
o
m
p
o
u
n
d
s
(
m
g

G
A
E
/
1
0
0

g

F
M
)

Irradiation dose (kGy)
0
A
a
0
2
4
6
8
10
12
14
b
a
b
a
B
D
P
P
H

r
a
d
i
c
a
l

s
c
a
v
e
n
g
i
n
g

a
b
i
l
i
t
y
(
m
M

T
E
A
C
/
g

F
M
)
Irradiation dose (kGy)
2 1 0.4 0
0
1
2
3
4
5
b
ab
b
a
C
A
B
T
S

r
a
d
i
c
a
l

s
c
a
v
e
n
g
i
n
g

a
b
i
l
i
t
y
(
m
M

T
E
A
C
/
g

F
M
)
Irradiation dose (kGy)
2 1 0.4 0
Fig. 1. Total phenolics content (A) and antioxidant capacity tested by DPPH (B),
ABTS (C) assays for gamma-irradiated pomegranate fruit (n = 3). Values with the
same letter for each dose are not signicantly different at p < 0.05.
316 H.M. Shahbaz et al. / Food Chemistry 145 (2014) 312318
irradiation may induce undesirable chemical changes, such as the
accumulation of volatile sulphur compounds including malondial-
dehyde, formaldehyde, and tetrahydrofuran which ultimately
leads to off-avour in fruit juices (Fan, Niemira, & Thayer, 2004).
Juice from the untreated fruits as well as from the irradiated
fruits were offered to the panelists to rate their preference. Table 4
shows the average and standard deviation values of the sensory
attribute ratings for pomegranate fruit juice. In general, the juice
from low dose (0.4 kGy and 1 kGy) treated fruits were liked more
among the panelists compared to the juice from the control and
high dose (2 kGy) treated fruits. The irradiated fruit juice samples
showed greater variance in the hedonic value at different dose lev-
els. The maximum average score was attained with the low dose
(0.4 kGy) treated fruit juice in respect to all sensory parameters.
Moreover, none of the samples were in the unacceptable hedonic
range (<2.5).
Irradiation induced a signicant impact on the colour of all the
juice samples similar to Hunter colour results. Noticeable changes
were observed and the colour of the juice from irradiated fruits
(0.4 kGy and 1 kGy) was more liked by the judges. Similar to our
results, Boylston et al. (2002) found that the colour of 0.75 kGy
irradiated rambutans and oranges were more intense than the con-
trol and preferred by panelists on the visual scale. Preferences for
overall taste were understood to increase with irradiation dose.
The total soluble solids and titratable acidity of the fruit juice did
not differ signicantly from irradiation (Table 1) corresponding
to all the juice samples also equally scored for their sweetness
and sourness values. The sweetness was the most liked attribute
by the panelists for all types of juices. This aspect also indicated
that sweet varieties of pomegranate fruit are liked among consum-
ers for fresh consumption as juice. The avour and aroma were
among the minimally affected sensory aspects as irradiation in-
duced no off-avors. Yu et al. (1995) also reported that sensory
panelists perceived no difference in the avour of strawberries
irradiated at 2 kGy. Irradiation enhanced the overall acceptability
of pomegranate fruit juice up to a 1 kGy dose.
In comparison to other studies, Kim and Yook (2009) found that
irradiation positively contributed to improve the sensory quality in
kiwifruits. Panelists showed a strong preference for irradiated
kiwifruits fruits (1, 2 and 3 kGy) in sweetness, overall taste and
overall acceptability. El-Samahy et al. (2000) also reported that
mangoes irradiated up 1.0 kGy were more acceptable to the sen-
sory panelists for their organoleptic properties.
Corresponding with the ndings of McDonald et al. (2012)
where consumers rated the overall acceptability of irradiated pea-
ches (up to 0.90 kGy) higher than that of the untreated peaches.
Mitchell et al. (1992) revealed that the sensory acceptability of or-
ange juice prepared from irradiated orange (0.6 kGy) fruit reduced
signicantly. Furthermore, Moreno, Castell-Perez, Gomes, Da Silva,
and Moreira (2007) stated that blueberries exposed to 3.2 kGy e-
beam irradiation were scored unacceptable by sensory panelists.
The sensory evaluation provides information about the attri-
butes of a product from consumers perspectives which determines
its acceptance. It is signicant to develop a relationship between
the physical and chemical composition of a product and its sensory
attributes, as well as between sensory perceptions and acceptabil-
ity for consumers (Escribano et al., 2010).
3.8. Correlation analysis
Correlation analysis of the data indicated (Table 3) that there is
a strong positive correlation between the antioxidant activity,
determined by the DPPH and ABTS assays, and total phenolic
(r = 0.8882 and r = 0.7700) and anthocyanins contents (r = 0.7671
and r = 0.5659). However, the correlation was higher for the DPPH
radial scavenging assay compared to the ABTS antioxidant assay.
These compounds are primarily responsible for the antioxidant
activity of pomegranate fruit juice. Similarly, total anthocyanins
content showed a signicant positive correlation with the total
phenolics content (r = 0.7375). A similar correlation was found by
Zaouay et al. (2012) between the total phenolics content and the
antioxidant capacity for 15 Spanish pomegranate cultivars. The
ndings also corroborates with the results of Mena et al. (2011)
who reported a linear correlation between total anthocyanins con-
tent and antioxidant activity assays. This is an indication that the
total anthocyanin compounds signicantly contribute to the anti-
oxidant activity in pomegranate fruit juice. Besides, there was a
signicant negative correlation between pH and titratable acidity.
4. Conclusions
Variability in the chemical composition and sensory proles of
the pomegranate fruit juice was observed at different dose levels
(0.4, 1 and 2 kGy). Irradiation improved the sensory prole and a
stronger preference was shown by the panelists for juice from
the 0.4 kGy and 1 kGy treated fruits. The chemical, sensory and
nutrient qualities of the juice from the fruits were most affected
by the higher 2 kGy treatment. Total phenolic compounds and
Table 3
Correlation coefcients (r) of total anthocyanins content (TAC), titratable acidity (TA), pH, reducing sugars (RS), total sugars (TS), total soluble solids (TSS), total phenolics content
(TP), DPPH and ABTS assays for antioxidant activity from juice of gamma-irradiated pomegranate fruit.
TAC TA pH RS TS TSS TP DPPH ABTS
TAC 1.0000
TA 0.6434 1.0000
pH 0.6749 0.9842 1.0000
RS 0.6038 0.9952 0.9684 1.0000
TS 0.8284 0.8835 0.8889 0.8592 1.0000
TSS 0.0658 0.0085 0.0143 0.0040 0.0000 1.0000
TP 0.8572 0.7375 0.7729 0.7104 0.9304 0.0178 1.0000
DPPH 0.8788 0.7671 0.7957 0.7545 0.8938 0.0274 0.8882 1.0000
ABTS 0.7686 0.5659 0.6590 0.5093 0.7496 0.2005 0.7700 0.7506 1.0000
Table 4
Sensory evaluation of juice from gamma-irradiated pomegranate fruit (n = 3).
Parameter Irradiation dose (kGy)
0 0.4 1 2
Color 3.70 1.02
ab
4.22 0.90
a
3.78 1.00
ab
3.26 0.92
b
Flavor 3.52 0.90
bc
4.09 0.85
a
3.87 0.92
ab
3.26 0.62
c
Aroma 3.30 0.76
a
3.43 0.59
a
3.48 0.67
a
3.35 0.57
a
Sweetness 4.13 0.69
a
4.09 0.51
a
3.96 0.47
a
4.00 0.43
a
Sourness 3.13 0.76
a
3.26 1.01
a
3.17 1.03
a
3.04 0.93
a
Overall taste 3.83 1.07
a
3.98 0.68
a
3.89 1.02
a
3.09 0.79
b
Overall
acceptance
3.43 0.66
b
3.98 0.61
a
3.83 0.58
a
3.13 0.76
b
Values with the same superscript letters (ac) in a row are not signicantly dif-
ferent at p < 0.05.
5 = Most like; 4 = good; 3 = fair; 2 = dislike a little; 1 = most dislike.
H.M. Shahbaz et al. / Food Chemistry 145 (2014) 312318 317
antioxidant assays were not signicantly affected up to the lower
0.4 kGy dose. Furthermore, there is a strong positive correlation
between the antioxidant activity, total phenolics and anthocya-
nins. On the basis of results, irradiation up to 1 kGy can be adopted
as a quarantine disinfestation treatment against disinfestation of
pests in pomegranate fruits. These research-oriented scientic
facts about irradiated pomegranate can help to boost the interna-
tional marketing and consumer acceptability of the fruit.
Acknowledgements
This research was supported by Export Promotion Technology
Development Program, Ministry of Agriculture, Food and Rural Af-
fairs, South Korea.
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