ChemicalEngineering&Processing:Pro

cessIntensification137(2019)1–11

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Chemical Engineering & Processing: Process Intensification

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Microwave-assisted extraction of phenolics from pomegranate peels:

Optimization, kinetics, and comparison with ultrasounds extraction
Kyriakos Kaderides, Lygeri Papaoikonomou, Melania Serafim, Athanasia M. Goula⁎
Department of Food Science and Technology, School of Agriculture, Forestry and Natural Environment, Aristotle University, 541 24, Thessaloniki, Greece

ART ICLEINFO
ABS TR ACT

Keywords:
Recently, there has been much interest in the phenolics of pomegranate peels, because of their health benefits,
Antioxidant activity
Microwave extraction such as antioxidant activity. Thus, the objective of this work is to optimize an efficient alternative and “green”
Phenolics technique for the extraction of phenolics from pomegranate peels. Microwave-assisted method was found to
Pomegranate peel increase extraction yield, but mainly to shorten the treatment time by over 60 times compared to conventional
Punicalagin extraction methods. The effects of solvent type, solvent/solid ratio, and microwave power on the yield of
Ultrasound extraction phenolics extraction were studied. The optimum operating conditions were found to be: solvent type, 50%
aqueous ethanol; solvent/solid ratio, 60/1 mL/g; power, 600 W. The efficiency of the proposed extraction pro-
tocol was compared with that of the ultrasound-assisted extraction, another “green” extraction approach, studied
in our previous work. The microwaves method had a yield about 1.7 times higher obtained in a shorter process
time (4 min) in comparison to ultrasound-assisted extraction (10 min). These differences were attributed to the
intense cell destruction of microwave-treated plant material observed by SEM analysis. The obtained extract
presented a high antioxidant activity (radical scavenging activity of 94.91%), due to the high content of puni-
calagin (143.64 mg/g dry matter) measured by HPLC analysis, that was equivalent to that of the ultrasounds
extract (radical scavenging activity, 94.77%; punicalagin content, 138.8 mg/g dry matter).

1. Introduction (e.g., as ingredient in functional foods). For example, PPE afforded

Pomegranate fruit (Punica granatum L.) is composed of three parts:
the seeds, the juice, and the peels. Pomegranate peels constitute ap-
proximately 40% of the whole fruit and are rich in phenolics com-
pounds such as ellagitannins, punicalagin, and punicalin [1]. Pome-
granate peels have been traditionally valorized as animal feed and
only small quantities are used in medicine. However, most of them are
di- rectly disposed as waste.
Several studies have reported the in vitro bioactivity of pome-
granate peel extracts including antioxidant, antitumor, anti-in-
flammatory, and antiproliferative properties. Kanatt et al. [2] in-
vestigated the antioxidant and antimicrobial potential of pomegranate
peel extract (PPE) and concluded that the efficacy of PPE in
scavenging hydroxyl and superoxide anion radical was very high. In
addition, the extract had good reducing power and iron chelation
capacity and showed good antimicrobial activity against
Staphylococcus aureus and Bacillus cereus having minimum inhibitory
concentration of 0.01%. Pseudomonas could be inhibited at a higher
concentration of 0.1% while it was ineffective against Escherichia coli
and S. typhimurium. Thus, PPE could potentially be included in several
industries due to its versatility
enhanced antioxidant activity by inhibiting lipid oxidation in cooked
chicken patties [3], whereas Kanatt et al. [2] found that addition of PPE
to popular chicken meat products enhanced their shelf life by 2–3
weeks during chilled storage. Pomegranate peel extract at concentra-
tions of 800–850 ppm in sunflower oil [4] and 200–1000 ppm in fish oil
[5] had stabilization efficiency comparable to conventional synthetic
antioxidants, i.e. BHT at its legal limit, whereas Kumudavally et al. [6]
and Devatkal et al. [7] concluded that PPE significantly increases the
stability of beef and goat meat products against lipid peroxidation.
According to Cam et al. [8], incorporation of pomegranate peel phe-
nolics into ice cream resulted in sharp improvements in antioxidant
and antidiabetic activities as well as the phenolic content of ice creams.
Addition of PPE to jams [9], juices and wines [10] increased their
phenolic, flavonoids, and thiol concentration with a significant im-
provement of the free radical scavenging and product stability
features. Recently, Kaderides et al. [11] incorporated pomegranate
peel extract in hazelnut paste and reported an inhibition of lipid
oxidation with reduced formation of peroxides.
Pomegranate peel phenolics can be extracted with conventional
systems such as Soxhlet and maceration. Shortcomings of existing

⁎
Corresponding author.
E-mail address: athgou@agro.auth.gr (A.M. Goula).

https://doi.org/10.1016/j.cep.2019.01.006
Received 22 November 2018; Received in revised form 3 January 2019; Accepted 10 January 2019
Availableonline24January2019
0255-2701/©2019ElsevierB.V.Allrightsreserved.
K. Kaderides et al. ChemicalEngineering&Processing:Proc
essIntensification137(2019)1–11

conventional extraction technologies, like increased consumption of dried

energy, high extraction time, possible degradation of bioactive com-
pounds, and high consumption of harmful chemicals, have forced the
food and chemical industries to find new separation “green’’ techni-
ques, which typically use less solvent and energy, such as ultrasound-
assisted extraction [11–14] and microwave-assisted extraction [15–17].
Microwave assisted extraction (MAE) is one of the most employed
alternative extraction methods. Among the advantages reported for
MAE are its lower extraction times and solvent consumption. The
process acceleration and the high extraction yield may be the result of a
synergistic combination of two transport phenomena: heat and mass
gradients working in the same direction [18]. According to Zheng et al.
[19], MAE is characterized by disruption of cell structure caused by the
penetrating volume heating from microwave irradiation. The diffusion
capacities of target components may be improved by the increment of
solvent temperature under microwave heating. So far, microwave
treatment has been widely employed to extract phenolics from different
plant materials including citrus peels [20], peanut skins [21], grape
seeds [22], tomatoes [23], blueberries [19], spent filter coffee [24],
chokeberries [25], and walnut leaves [26].
Therefore, MAE is a potential method for the extraction of
phenolics from pomegranate peels. According to Zheng et al. [19], the
optimi- zation of extraction parameters is helpful to systematically
understand the interaction between independent factors and obtain
the optimal parameter combination for optimum extraction. In
addition, the MAE efficiency depends on several variables which may
not be generalized for all plant materials due to the diverse nature of
existing bioactive phytochemicals, being necessary to select and
optimize the processing conditions as a function of the used matrix
and taking into account the desired responses [27]. To our best
knowledge, as far as pomegranate peel is concerned, only Zheng et al.
[16] and Huang et al. [17] em- ployed MAE to extract phenolic
compounds with water and flavonoids with aqueous ethanol,
respectively.
According to Shirsath et al. [28], mathematical modelling can help
in the design, optimization, and control of the extraction process, as
well as provide useful information for scale up of the equipment. The
typical kinetic models of extraction include unsteady diffusion, Fick’s
law of diffusion, film theory, and empirical models. However, no in-
formation is available for kinetic modeling of microwave-assisted ex-
traction of phenolics from pomegranate peels.
Therefore, the objective of this study is to develop an efficient al-
ternative and “green” technique for the extraction of phenolics from
pomegranate peels. The operating parameters are optimized and the
mechanism of the enhanced extraction is discussed by observing cell
destruction of plant material. The efficiency of the proposed extraction
protocol is compared with that of another “green” extraction approach.
The content of the major phenolic compounds and the antioxidant ac-
tivity are also evaluated. In addition, a mathematical model to describe
the kinetic mechanisms of polyphenols extraction is developed,
whereas the dependence of the best descriptive model constants on
extraction variables is expressed by an appropriate type of model.
These results would offer scientific reference for promoting the
valorization of pomegranate peels.

2. Materials and methods

2.1. Standards and reagents

HPLC-grade acetonitrile was from Fisher Scientific (Lisbon,

Portugal). The phenolic compound standards were from Sigma-
Aldrich (St. Louis, MO, USA). All the other chemicals were of
analytical grade.

2.2. Pomegranate peels

Pomegranate peels as by-product of fruit juice industry were pro-

vided from a local producer (Rodi Hellas, Greece). The peels were

2
K. Kaderides et al. ChemicalEngineering&Processing:Proc
at 40 °C for 48 h and kept at −30 °C until use. The peels were ground essIntensification137(2019)1–11

in a laboratory mill (Type A10, Janke and Kunkel, IKA Labortechnik,

Germany) immediately prior to extraction. The particle size
distribution of the milled pomace was determined by a sieve shaker
(Retax, Labor Siebmaschine, Type LS10, Nr 4082, Germany). The
particle mean dia- meter was about 0.1 mm.

2.3. Microwave-assisted extraction

Microwave-assisted extraction was performed with a Multiwave

closed microwave system (B30MC030A, AntonPaar, Graz, Austria),
equipped with standard configuration with rotor for 6 sample vessels
type H (50 mL quartz tubes; operating pressure 75 bars). A sample of
powdered pomegranate peels was mixed with 20 mL solvent to produce
different solvent/peel ratios. The extracts were collected at 0.5, 1, 2,
3, 4, 5, 10, and 15 min. The resulting extracts were evaporated using a
rotary evaporator (Rotovapor R114, Waterbath B480, Büchi, Flawil,
Switzerland). The variation of extraction yield during the extraction
process was studied with various (i) microwave powers (P) (100, 173,
350, 527, and 600 W) and (ii) solvent/peels ratios (L/S) (10.0, 17.3,
35.0, 52.7, and 60.0 mL/g). A central composite design was applied to
determine the effects of the above parameters on extraction yield.
Thirteen experimental runs were carried out and each one was con-
ducted with five different solvents: water, 50 and 70% aqueous
ethanol, and 50 and 70% aqueous methanol.
Extraction yield, Y (%), was defined as the percent ratio of the
total weight of gallic acid equivalents extracted to the dry peels
sample weight.

2.4. Determination of total phenolics content and antioxidant activity

The total phenolic content of extracts was determined according

to the Folin-Ciocalteu method [29]. Briefly, 0.2 mL of the extract was
added to a 25 mL volumetric flask and additional distilled water was
added to make a final volume of 10 mL. A reagent blank was prepared
using distilled water. Folin-Ciocalteu phenol reagent (0.5 mL) was
added to the mixture and shaken vigorously. After 5 min, 5 mL of 5%
Na2CO3 solution was added with mixing. The solution was
immediately diluted to 25 mL with distilled water, mixed thoroughly
and then al- lowed to stand for 90 min. The absorbance was measured at
750 nm versus the prepared blank. The total phenolic content of the
sample was expressed as gallic acid equivalents (GAE) mg/mL.
Antioxidant activity was determined using DPPH, which is a
stable free radical and is widely used to assess the radical scavenging
activity of antioxidant components. This method is based on the
reduction of DPPH in methanol solution in the presence of a
hydrogen donating antioxidant due to the formation of the non
radical form DPPH-H [30]. The transformation results in a color
change from purple to yellow, which is measured
spectrophotometrically at 517 nm. The reaction mixture (3.0 ml)
consists of 1.0 ml of DPPH in methanol (0.3 mM),
1.0 ml of extract, and 1.0 ml of methanol. The percentage of inhibition
can be calculated using the Eq. (1).

Inhibition (%) = ((A0–A1)/A0) × 100 (1)

where A0 is the absorbance of the control and A1 is the absorbance of

the sample.

2.5. Analysis of phenolic compounds

The samples were analyzed using a HPLC system equipped with two
Marathon IV pumps (Spark NL., The Netherlands), a Timberline TL-
50 temperature controller (Indiana, U.S.A.), a UV–vis (UV 6000 LP de-
tector -Spectra SYSTEM, Finningan Mat, Thermo Separation
Products, San Jose, U.S.A) and a hypersil C18 reversed-phase
column (250 × 4.6 mm I.D., 5 μm particle size). The mobile phase
was a

3
mixture of water/acetic acid (95:5) (A) and acetonitrile (B). Samples
were analyzed using a gradient program developed by Kaderides et
al. [31]. Analyses were conducted at constant temperature of 30 °C using a
flow rate of 1 mL/min and a sample injection volume of 10 μL. Double
online detection was carried with a diode array detector operating at
254 and 280 nm as preferred wavelengths and the target phenolic
compounds were identified according to their UV spectra and
retention time. For the quantitative analysis, a baseline to valley
integration with
baseline projection mode was used to calculate peak areas and external
standards were used for quantification. The results were expressed in
mg per g of dry weight.
2.6. Scanning electron micrographs (SEM)
A Quanta-200 environmental scanning electron microscope system
(FEI Company, USA) was used to study the structural changes in po-
megranate peels texture after the extraction process. The samples were
fixed on a specimen holder with an aluminum tape and sputtered with
a thin layer of gold prior to examination under high vacuum condition
at an accelerating voltage of 15 kV at 2000× magnification level.
2.7. Kinetic models
In the present study, three different mathematical approaches were
selected for the kinetic modelling of extraction of polyphenols, namely,
the Peleg’s model, the first-order kinetic model, and the second-order
kinetic model. These three models were compared by fitting the ex-
perimental data obtained from extraction into each model.
2.7.1. Peleg’s model
Peleg’s equation [Eq. (2)] is a non-exponential empirical equation
proposed as [32]:
t A p value less than 0.05 was considered to be statistically
Ct =
K1 + K2⋅t (2)
where Ct is the concentration of polyphenols in the extract (mg/mL) at
time t (min), K1 is Peleg’s rate constant (min mL/mg), and K2 is Peleg’s
capacity constant (mL/mg). Peleg’s K1 parameter refers to extraction
rate (B0, mg/min mL) at the very beginning (t = t0) according to Eq.
(3):
0= 1
B K1 (3)
Peleg’s capacity constant K2 refers to the maximum extraction
yield, that is, the equilibrium concentration of total extracted analyte
when t
→ ∞ (Ceq, mg/mL), as indicated in Eq. (4).
eq = 1
C K2 (4)
2.7.2. First-order kinetic model
According to Poojary and Passamonti [32], when we plot the con-
centration of extracted polyphenols in the matrix against time, the
curve which describes the extraction kinetics usually shows an ex-
ponential decay. The most usual form of the first-order kinetic model
is presented in Eq. (5).
Ct = Ceq (1 − e−KA⋅t )
(5)
where KA is the overall extraction rate constant (1/min).
2.7.3. Second order kinetic model
The solid-liquid extraction process can be considered as the reverse
of an adsorption operation, therefore the bases of the adsorption
kinetic equations can be applied to solid-liquid extraction and the
dCt
= KB⋅(Ceq − Ct )2
d (6)
t
where KB is the second-order extraction rate constant (mL/mg min).
The integrated rate law for a second-order extraction under the
boundary conditions t = 0 to t and Ct =0 to Ct, can be written as an Eq.
(7):
2
KB⋅t⋅Ceq
Ct =
1 + KB⋅t⋅Ceq (7)
where h is the initial extraction rate (mg/mL min) when t approaches 0:
h = KB⋅Ce2 (8)
q
2.8. Statistical analysis
The parameters of the kinetic models were estimated by nonlinear
regression. To evaluate the goodness of fit of each model, two criteria
were used: the coefficient of determination, R2, which is the relative
variance explained by the model with respect to the total variance, and
the standard error of the estimate, SEE (Eq. (9)), which is a measure of
the accuracy of predictions.
SEE ∑ (Ct,exp − Ct,pred )2
=
N (9)
where Ct,exp is the experimental value of polyphenols concentration in
the extract at time t, Ct,pred is the predicted concentration, and N is the
number of observations.
The effect of process variables on extraction yield and on kinetic
parameters were analyzed using the statistical software MINITAB
(Release 13.32). To identify the significance of the effects and inter-
actions between them, analysis of variance (ANOVA) was performed.
significant.
Regression analysis was used to fit full second order polynomials, re-
duced second order polynomials, and linear models to the data of each
of the variables evaluated (response variables). F values for all reduced
and linear models with an R2 > 0.70 were calculated to determine if
the models could be used in place of full second order polynomials to
predict the response of a variable to the independent variables.
3. Results and discussion
3.1. Effect of process variables on microwave-assisted extraction yield
The extraction yield was time-dependent and increased with ex-
tended irritation times from 0.5 to 4 min and then slightly decreased
(Fig. 1). Thus, the efficient extraction period for achieving maximum
yield of pomegranate peel phenolics was about 4 min. According to
Veggi, Martinez, and Meireles [18], the extraction process takes place
in three different steps: an equilibrium phase where the phenomena of
solubilization and partition intervene, in which the substrate is re-
moved from the outer surface of the particle at an approximately con-
stant velocity. Then, this stage is followed by an intermediary
transition phase to diffusion, in which mass transfer by convection and
diffusion prevails. In the last phase, the extraction rate is low and the
solute must overcome the interactions that bind it to the matrix and
diffuse into the
extracting solvent. During microwave-assisted extraction of pome-
granate peel phenolics, the absorption of microwave energy in the ex-
traction system promoted the thermal accumulation of the extraction
solution resulting in the dissolution of phenolics into the solution until
4 min [34]. The same behavior was observed by Xie et al. [35] and Wu
second-order law was found to give the best fits for the extraction rate
[33]. The general second-order model can be written as [13]:
et al. [36] for extraction of flavonoids from Cyclocarya paliurus Iljins-
kaja leaves and phenolics from potatoes, respectively. Huang et al.
[17] and Alara et al. [37] reported an initially increase in extraction
yield with increasing irritation time reaching a maximum value,
during ex- traction of phenolics from pomegranate peels and
Vernonia amygdalina
Fig. 1. Main effects plot illustrating the effect of extraction time (t, min), microwave power (P, W), and solvent-to-peels ratio (L/S, mL/g) on the extraction yield (Y,
mg GAE/g dry peel).

leaves. However, in all these studies the excessive time exposure in the
methyl radical present in methanol, resulting in a lower solvation of
microwave field decreased the extraction yield due to a possible de-
antioxidant molecules [39]. However, in the case of microwave ex-
gradation of phenolic compounds [37].
traction, the loss tangent is the ratio of the dielectric loss to the di-
The most important factor that affects MAE is solvent selection that
electric constant, a dielectric parameter which can be used as an index
depends on the solubility of the compounds of interest, solvent pene-
of the efficiency with which electromagnetic radiation is converted to
tration and its interaction with the sample matrix and its dielectric
heat [40]. The loss tangent of ethanol and methanol at 2.45 GHz and
constant, and the mass transfer kinetics of the process [18]. In this
30 °C is 0.898 and 0.522, respectively [41], and of water at 2.45 GHz
work, the ability of different solvents in extracting phenolic
and 25 °C is 0.123 [42]. This means that water is considered a medium
compounds from pomegranate peels was compared. Fig. 2 is the main
microwave absorber, methanol is a good absorber, while ethanol is
effects plot for solvent type, which was found to have a
considered a strong microwave absorber. Hence, ethanol has higher
significant effect (p < 0.05) on total phenolic yield. Extraction yield
ability than methanol to absorb microwaves and convert them to heat,
increased in the following order of solvents: 70% methanol < 50%
promoting both the rapid rupture of the plant material and the in-
methanol < water < 70% ethanol < 50% ethanol.
creased diffusion of the targeted compounds [40].
Ethanol-water mixtures seem to be the most suitable solvents for
In addition, it was found that a binary solvent is more efficient for
the extraction. Kaderides, Goula, and Adamopoloulos [11] reported
extraction of phenolics than a mono-solvent one [43]. Addition of
that the extraction yield increases with an increase in solvent polarity
water to a solvent usually creates a more polar medium, which
and a decrease in viscosity and the higher the molecular weight of the
facilitates the extraction of polyphenols as suggested by Spigno et al.
solvent, the lower the polarity, which allows other substances of about
[44]. By in- creasing the proportion of water, the polarity of solvent
the same molecular weight to be easily extracted. This can be
also increases and the system is able to extract phenolic substances
correlated to “like dissolve like” or “polarity versus polarity” principle.
from both ends of the polarity (highest polarity substances and low
Ethanol, methanol, and water have a relative polarity of 0.654, 0.762,
polarity substances), as well as those of moderate polarity. Several
and 1.0, respectively [38]. Methanol is considered more efficient
researchers reported that an increase of ethanol concentration in a
solvent for extraction of antioxidant compounds than ethanol even if
binary ethanol-water solvent results in the addition of the polarity
their polarities are similar. This may be due to the low solvation
molecular to close that in phenolic compounds. Thus, the phenolics
provided by ethanol, probably because of the presence of the ethyl
solubility may be improved with the ethanol concentration based on
radical that is longer than the
the “polarity versus polarity” principle. Zheng et al. [19] explained that
ethanol plays a major role to rupture the hydrogen bonds and
hydrophobic bonds existing between pheno- lics–proteins and
phenolics–cellulose in the water–ethanol system. However, in the
present work, 50% ethanol was found more efficient than 70% ethanol.
This finding may be associated to the fact that both the dielectric
constant and dielectric loss factor of water are higher than those of
ethanol and under the same irradiated intensity of microwave output
power, water may absorb more thermal energy than ethanol to drive
target component to diffuse [19]. A similar trend was reported by
Zheng et al. [19], who concluded that the more appropriate ethanol
concentration for microwave-assisted extraction of anthocyanin from
blueberry is around 60%, and Taha et al. [45], who studied the effect of
ethanol concentration (ranged from 50 to 80%) on the total content of
phenolic compounds in sunflower meal extracts, found that 60%
ethanol was the most efficient.
Figs. 1 and 3 presents the effects of microwave power (P) and sol-
Fig. 2. Main effect of solvent type on the extraction yield (Y, mg GAE/g dry
vent/peel ratio (L/S) on extraction yield. As the liquid-to-solid ratio
peel) obtained at 4 min.

Fig. 3. Contour plots illustrating the effect of microwave power (P, W) and
solvent-to-peels ratio (L/S, mL/g) on the extraction yield (Y, mg GAE/g dry
peel) obtained at 4 min using 50% aqueous ethanol.
increases from 10.0/1 to 52.7/1 mL/g, the extraction yield is sig-
nificantly enhanced. In general, a higher ratio of solvent to raw
material results in a larger concentration gradient during the diffusion
from the solid into the solution and a higher extraction yield. In
addition, the increase of solvent/solid ratio leads to excessive swelling
of the plant material increasing the contact surface area between the
material and the solvent [34]. However, with further increase in
liquid/solid ratio, a decrease in yield was observed. Shirsath et al. [28],
who studied the extraction of curcumin from Curcuma amada,
concluded that if the so- lution is very dilute, an increased quantity of
solvent would not lead to a sufficient increase in the concentration
gradient and the increase in extraction yield would be limited. A large
solvent volume would de- crease the microwave adsorption of
material, since more energy is needed to be absorbed by the solvent
[17]. According to Alara et al. [37], a very high solvent/solid ratio may
result to insufficient energy in facilitating the cell wall breakage for
effective leaching out of the phenolic compounds. A similar observation
was reported by Galan et al.
[40] and Huang et al. [17], who found the maximum extraction yield of
phenolics from Citrus sinensis peels and pomegranate peels at a
liquid/ solid ratio of 30/1 and 40/1 mL/g, respectively. Also, using a
large amount of solvent is not considered to be cost-effective due to
high operating cost of solvents and energy consumption required
during the subsequent separation [28]. However, Zheng et al. [16]
reported that the effect of liquid/solid ratio on recovery of phenolics
from pome- granate peels was not significant. Thus, the optimum
value of solvent/ solid ratio is specific to the system under
investigation and needs to be established experimentally.
As far as the effect of microwave power (P) is concerned, the ex-
traction yield generally increased with increasing the power from 173
to 600 W, whereas an opposite trend was observed when power in-
creased from 100 to 173 W. It must be underlined that the extraction
temperature, which increases with microwave power, could not be
adjusted in the microwave system used in the present work. The first
effect may be attributed to the fact that increasing the microwave
power, more electromagnetic energy was transferred on the molecules
by ionic conduction and dipole rotations resulting in rapidly heating
of the extraction system [46]. At higher temperatures, the solubility
and diffusion coefficients of the phenolic compounds to be extracted
in- crease and the viscosity of the solvent decreases, thus facilitating its
passage through the solid substrate mass. Moreover, the microwave
irradiation accelerates cell rupture by sudden internal pressure
increase inside the cells of plant sample, which promotes the
destruction of sample surface and in turns the exudation of phenolic
compounds within the plant cells into the surrounding solvent [47].
According to Liew et al. [48], some researchers prefer the approach of
low micro- wave power with longer extraction time as high power
may reduce purity. However, this approach may cause overheating
and reduce the
nutrient content of the extracted compounds. However, at power
lower than 173 W, a decrease of extraction yield was observed with
increasing the power. According to Maran et al. [49], this effect may be
due to the fact that high irradiation power offered redundant energy
to the solvent and material drastically disturbing molecular
interactions.
According to the ANOVA analysis of the optimization study, the
liquid/solid ratio and its square, both with a probability value of 0.001,
influenced most significantly the extraction yield. The regression
coefficients were calculated and the data was fitted to a second-order
polynomial equation. The response extraction yield (Y) can be ex-
pressed in terms of the regression Eq. (10):
Y = 140.62 − 0.008⋅P + 0.996⋅L/S + 0.002⋅(L/S)2 (10)
2
The coefficient of determination, R , was 0.912, indicating that 91.2%
of the total variability in the response could be explained by the
specific model, whereas an adjusted R2 value of 0.891 revealed that
there was a good agreement between experimental and predicted
values of ex- traction yield. The model F-value was 3.15,
corresponding to a p-value of less than 0.05, which, along with the p-
value for lack of fit (0.188), indicated that the model fits the
experimental data accurately.
The desirability profile for optimum yield indicated that the max-
imum desirability level of 1.0 (on a scale of 0–1) can be achieved with a
microwave power of 600 W, a solvent/solid ratio of 60/1 mL/g, and
50% aqueous ethanol as solvent at an extraction time of 4 min. Under
these optimized conditions, the predicted value for extraction yield
was
202.8 mg GAE/g dry peels, whereas the observed experimental value
was found 199.4 mg GAE/g dry peel. A similar value was reported by
Zheng et al. [16], who found the optimum extraction yield of total
phenolics from pomegranate peel at 214.5 mg GAE/g dry peel.
3.2. Microwave-assisted extraction kinetics
Different kinetic models were employed to fit with experimental
data. The highest values of R2 and lowest values of SEE were chosen
for goodness of fit. Table 1 presents the R2 and SEE values for the
kinetic models. Among these, the Peleg’s model fits the best with
experimental values showing the highest average values of R2 (0.924–
0.991) and the lowest average values of SEE (0.889–1.322). The
applicability of Peleg’s equation on food materials has been
demonstrated extensively [32].
The kinetic parameters of the Peleg’s model at different extraction
conditions are presented in Fig. 4. It should be noted that a lower K1
value in Eq. (2) implies a faster rate of the process, whilst the lower K2
value indicates maximum yield [50]. According to Ji et al. [51], the
extraction rate is controlled not only by mass transfer resistance in the
liquid film around solute particles, but also by intra-particle diffusion;
hence when the mass transfer resistance in the liquid film is reduced,
the extraction rate is controlled only by intra-particle diffusion. As it
can be seen, the kinetic parameters generally increased with an
increase in liquid-to-solid ratio. Higher liquid to solid ratio ensures
homogenous mixing and allows penetration of solvent into deep
interior parts [52]. However, the capacity and the rate decreased with
an increase in L/S. This effect may be attributed to the fact that for
microwave irradiation the temperature rise is higher at lower solvent-
to-feed ratio owing to the lower volume of solvent [53]. As far as
microwave power is con- cerned, power provides localized heating in
the sample, which acts as a driving force for MAE to destroy the plant
matrix so that the solute can diffuse out and dissolve in the solvent,
thus increasing the rate constant. However, for high solvent-to-solid
ratios, rapid rupture of the cell wall can take place at a higher
temperature when using higher power, and as a result impurities can
also be leached out into the solvent together with the desired solute
[18]. Thus, the constant K1 presents a maximum at a microwave power
around 400 W. As it can be seen in Fig. 4, the ex- traction capacity
slightly decreased with an increase in P. This finding may be
associated with a possible degradation of thermally sensitive
compounds at high microwave powers.
Fig. 5 presents the comparison of predicted and experimental
values
Table 1
Statistical parameters of the kinetic models for microwave-assisted extraction of polyphenols from pomegranate peels using 50% aqueous ethanol.
A/A Microwave power (P, W) Solvent/peel ratio (L/S, mL/g) Kinetic model

Peleg’s model First-order kinetic model Second-order kinetic model

R2 SEE R2 SEE R2 SEE

1 350 10.0 0.972 1.322 0.971 1.361 0.965 1.481

2 527 17.3 0.991 0.498 0.987 0.606 0.973 0.885
3 173 17.3 0.965 0.839 0.958 0.919 0.911 1.347
4 100 35.0 0.964 0.478 0.935 0.640 0.907 0.769
5 600 35.0 0.976 0.468 0.974 0.486 0.938 0.744
6 350 35.0 0.978 0.401 0.952 0.584 0.941 0.650
7 350 35.0 0.972 0.442 0.938 0.651 0.908 0.795
8 350 35.0 0.982 0.376 0.952 0.623 0.922 0.793
9 350 60.0 0.930 0.539 0.962 0.399 0.509 1.431
10 350 35.0 0.953 0.630 0.920 0.818 0.912 0.861
11 527 52.7 0.977 0.289 0.978 0.286 0.948 0.433
12 350 35.0 0.979 0.438 0.953 0.653 0.893 0.983
13 173 52.7 0.924 0.523 0.882 0.652 0.832 0.777
Average 0.966 0.557 0.951 0.668 0.889 0.919

Fig. 5. Experimental and predicted by the Peleg’s model values of polyphenols

concentration in the extract (C, mg/mL) at a microwave power of 350 W, a
solvent-to-peels ratio of 35.0/1 mL/g, and 50% aqueous ethanol as solvent.
Prediction Band is the region where 95% of the experimental data points are
expected to be and Confidence Band is the region where 95% of the regression
lines are expected to be.

Qu et al. [54], who studied the extraction of antioxidants from

Fig. 4. Contour plots illustrating the effect of microwave power (P, W) and
solvent-to-peels ratio (L/S, mL/g) on the kinetic parameters of the Peleg’s
model (K1 in min mL/mg and K2 in mL/mg).
of polyphenols concentration in the extract for the Peleg’s model at a
microwave power of 350 W, a liquid-to-solid ratio of 35.0/1 mL/g, and
50% aqueous ethanol as solvent. The figure presents also the
confidence (CB) and prediction (PB) bands for the polyphenols
concentration at the solvent phase against time. It is noted that similar
trends were observed
for all experiments. The PB is the region where 95% of the
experimental data points are expected to be and it is observed that all
obtained ob- servations fall within. Likewise, the CB is the region
where 95% of the regression lines are expected to be.
pomegranate marc, fitted the relationships between kinetic parameters
and processing factors (particle size, water/sample ratio, and tem-
perature) by linear, power or second-order polynomial functions.
However, they developed a different model for each factor. In this
work, multiple regression analysis was used to develop equations pre-
dicting the effect of all extraction factors simultaneously. The effect of
extraction variables on Peleg’s model constants can be expressed by
Eqs. (11) and (12).
K1 (×10−3) = 4.006 + 0.445⋅P − 5.469⋅L/S + 0.153⋅L/S2
− 0.003⋅P⋅L/S (R20.911) (11)
K2 (×10−3) = 42.467 + 0.166⋅P − 3.641⋅L/S + 0.018⋅L/S2
+ 0.001⋅P⋅L/S (R2 = 0.962) (12)
Substituting Eqs. (11) and (12) into Eq. (2), a kinetic model for
predicting polyphenols extraction from pomegranate peel can be ob-
tained. Even though the empirical models (11)–(12) cannot account for
the phenomena governing extraction processes, they could be used to
determine the effects of microwave power and liquid-to-solid ratio on
 Fig. 6. HPLC chromatogram of pomegranate peel extract at the optimum conditions of microwave- and ultrasound-assisted extraction (254 nm).
the phenolics extraction yield [33].
3.3. HPLC analysis
Polyphenol qualitative and quantitative determination of pome-
granate peel extract was performed by HPLC analysis. The identified
phenolic components were quantified with respect to reference sub-
stances to compare the content of the major individual phenolic com-
ponents in pomegranate peel extract. Generally, limited research has
been achieved with regard to the quantification of phenolic
compounds in pomegranate peel extracts using HPLC; furthermore, it
is difficult to compare results from different studies since different
extraction methods were used. In the present work, punicalagin was
determined as predominant in the extract at the optimum conditions of
microwave- assisted extraction (Fig. 6). This observation is similar to
that reported by other researchers. Aqil et al. [55] and Gullon et al. [56]
reported that punicalagin is the main bioactive compound present in
pomegranate peel. Similarly, Yan et al. [57] found that among the
phenolic
compounds of pomegranate peel, punicalagin present as a mixture of
60 and 240 min, respectively. Cam and Hisil [1] reported a maximum
two reversible anomers (a and b) was the dominant one, accounting
yield of 264 mg tannic acid equivalents/g dry matter at 15 min with the
for more than 90% of the phenolics in all of the tested cultivars (range,
pressurized water extraction method. Mushtaq et al. [71] proposed an
17.8–114.5 mg/g). The alpha and beta forms of pomegranate punica-
enzyme-assisted supercritical fluid extraction of phenolics form pome-
lagin are polyphenolic hydrolysable tannins and isomers of 2,3-(S)-
granate peels with an optimum yield of 301.53 mg GAE/g extract at
hexahydroxydiphenoyl-4,6-(S,S)-gallagyl- D-glucose [58].
85 min.
Punicalagin content (a + b) of the pomegranate peel extract at the
As far as new separation “green’’ techniques are concerned, ultra-
optimum conditions of microwave-assisted extraction was determined
sound- and microwave-assisted extraction are energy-saving technolo-
as 143.64 mg/g on dry matter basis. This punicalagin content is higher
gies that are increasingly employed in the extraction of natural
than the values reported by Lu et al. [59] (39.8–121.5 mg/g dry weight)
products as alternatives to traditional techniques of extraction. Pan et
for pomegranate peel extracts obtained by UAE for 30 min in 40%
al. [13] reported extraction yields, under the optimum conditions of
aqueous ethanol. Gullon et al. [56] applying ultrasound-assisted ex-
ultrasound extraction of phenolics from pomegranate peel with water,
traction with 50% aqueous ethanol determined a lower punicalagin
of 14.8 and 14.5% at extraction times of 6 and 8 min for continuous UAE
content of 16.67 mg/g dry weight. On the contrary, Kharchoufi et al.
and pulse UAE, respectively. Tabaraki et al. [14] achieved an
[60] using the normal stirring method, reported much higher values of
optimum yield of
punicalagin, respectively 193.78 mg/g for a methanolic extract and
91.98 mg GAE/g dry peels using 70% ethanol–water mixture as solvent
245.47 mg/g for an aqueous one. Such variations could be due to the
and extraction time of 30 min. In our previous work, Kaderides et al.
extraction conditions and solvents used and probably also to the
[11] studied the effects of various parameters on ultrasound-assisted
variability associated with the difference in the stage of maturity of
extraction yield of pomegranate peel phenolics and found the
cultivars and geographical area of origin [60]. According to Saad et al.
optimum conditions to be: solvent type, water; extraction time, 10 min;
[61], total polyphenol levels increase at the early stage of growth in
tem- perature, 34.7 °C; solvent/solid ratio, 32.2/1; amplitude level,
peel, but thereafter generally decrease during maturation.
39.8%; pulse duration/pulse interval ratio, 1.2/1. Under these
Yan et al. [57] found an ellagic acid content of 0.39–3.04 mg/g in
optimized con- ditions, the experimental value of extraction yield was
pomegranate peel extract and similarly, Seeram et al. [62] reported that
119.82 mg GAE/ g dry peels at an extraction time of 10 min. The
pomegranate peel contains major ellagitannins, such as punicalagin
obtained UAE extract was analysed in the present work for polyphenol
(80–85% w/w) and ellagic acid (1.3% w/w). Zhou et al. [63] and Cam
qualitative and quantitative determination, antioxidant activity, and
and Hisil [1] identified ellagic acid and the other ellagic acid deriva-
surface mor- phology.
tives, which were not determined in the present study. This may be
The microwave-assisted extraction method that was used in the
attributed to the smaller solubility of ellagic acid in aqueous ethanol
present work provided higher extraction efficiencies in comparison to
compared to methanol, which was used as an extraction solvent in the
ultrasound-assisted extraction (Table 2). The MAE of pomegranate
aforementioned studies. Panichayupakarananta et al. [64] investigated
peel phenolics had a maximum yield of 199.4 mg GAE/g dry peel at an
the influence of different methanol concentrations on the content of
ex- traction time of 4 min A similar trend was observed by Ince et al. [ 72]
ellagic acid in pomegranate peel extracts and concluded that 90% v/v
during the extraction of phenolic compounds from nettle by micro-
methanol gave significantly higher content of ellagic acid compared to
waves (24.64 mg GAE/g dry material) and ultrasounds (23.86 mg GAE/
other analyzed methanol percentages, whereas Rodrigues et al. [65]
g dry material). The solvent accessibility to the pomegranate peels
also showed that during UAE of jabuticaba tree, the best results for
must have been enhanced due to the action of microwaves, disrupting
ellagic acid were achieved at high ethanol concentration and high ex-
the peels matrix and allowing the penetration of solvent. Generally,
traction time. In addition, Elfalleh et al. [66] reported a gallic acid
ultra- sound waves and the cavitation effect they produce can alter
content varied from 1.09 to 1.31 mg/g in six varieties of Tunisian po-
biological materials, facilitating the release of extractable compounds
megranate peel after methanol extraction overnight. Generally, al-
and en- hancing mass transport by disrupting cell walls, whereas the
though the existence of gallic acid, caffeic acid, chlorogenic acid, p-
effec- tiveness of microwaves is attributed to its localized heating
coumaric acid, catechin, quercetin, and rutin in pomegranate peels
which in- creases the internal pressure of the cells and consequently
were reported in the literature, these polyphenols were not identified
ruptures them. Heat and mass transfer occur in the same direction
in the present work. Gullon et al. [56] reported that the type and con-
from inside to outside in MAE, accelerating the solubilisation of
centration of bioactive compounds in pomegranate peel extract
solutes. Meanwhile the heat that transferred from the outside to the
depend on several factors, such as cultivar analyzed, soil factors,
inside of samples in the ultrasounds method prolongs the extraction
environmental conditions, maturity stages and so on. According to
period needed for the so- lutes to diffuse out and become solubilised
Bruni [67], the proclivity to hydrolysis and in particular the different
in the solvent.
solubility in water of some ellagitannins according to their molecular
The raw material (RM) and the treated by MAE and UAE peels at the
weight, are factors that must be taken into account when comparing
optimum conditions were examined by scanning electron microscopy
phytochemical profiles of pomegranate peels in the literature.
(SEM) and their micrographs are shown in Fig. 7. The surface of the
Generally, a number of researchers have concluded that the solvent
peels sample before extraction was densely packed, whereas extraction
used, the extraction method, and the duration of the process may
produced structural changes in all samples. After UAE, no significant
severely affect both the amount of phenolics and their relative ratios.
changes could be found for the surfaces of samples except for few
creases and ruptures, which may be due to the cavitation effects of
3.4. Comparison between microwave- and ultrasound-assisted extraction ultrasounds [73]. On the contrary, in the case of MAE, the sudden
thermal stresses and the high localized pressures affect the cell
Several researchers extracted phenolic compounds of pomegranate structure of the cell causing intense creases and ruptures. Dahmoune
peel using different extraction methods (Soxhlet, normal stirring, et al. [74] confirmed in their work on the microwave extraction of
pressurized water extraction, supercritical fluid extraction) [11] and polyphenolic compounds from medicinal plants that the
reported much longer extraction times (up to 60 times) to achieve electromagnetic waves caused cell damage and rupture due to sudden
lower, similar or slightly higher extraction yields. Negi et al. [68] ex- temperature rise during microwave irradiation. Alara et al. [37] also
tracted phenolics from pomegranate peels using the Soxhlet method reported that microwave irradiation plays an important role in vegetal
with water for 4 h and reported a yield of 4.8%. Pan et al. [69] and cell wall breakage and this phenomenon might be due to the rotation
Wang et al. [70] used the normal stirring method with water and of liquids and the mo- lecular movement associated with a permanent
achieved a total phenolic yield of 119 and 82.6 mg GAE/g dry matter at dipole, resulting in rapid heating of the solvent. This structural change
allows easy entry of the solvent into cellular channels with high
extraction efficiency.
In addition, the lower total phenolics content obtained by the
 ultrasound-assisted method could be attributed to a possible degrada-
tion of the extract due to the generation of hydroxyl radicals by
acoustic cavitation during ultrasounds treatment in water [75].
Punicalagin content (mg/g dry matter)Antioxidant activity (radical scavenging
Vinatoru et al.
[76] reported that the presence of water vapor in the bubbles leads to a
homolytic splitting of the water molecules generating reactive HO and
hydrogen atoms during its collapse. The radicals formed then undergo
reactions to produce H2O2 and other active oxidising agents resulting
in possible phenolics degradation.
Punicalagin was found to be the major polyphenol in the
optimized pomegranate peel extract for both MAE and UAE, with a
content of
143.64 and 138.8 mg/g dry matter, respectively (Fig. 6). Various po-
tential biological and pharmacological activities, including anti-bac-
terial, anti-mutagenic, anti-carcinogenic, anti-inflammatory, hepato-
protective, and anti-genotoxic properties, have been attributed to the
presence of punicalagin, mostly due to its high antioxidant activity
[56]. Gil et al. [77] determined the antioxidant activity of individual
94.77

phenolic groups in pomegranate juice by the DPPH method and found

that punicalagin had about 1.5-, 6.5-, and 20-times higher antioxidant
activity than those of hydrolyzable tannins, anthocyanins, and ellagic
94.91

acids, respectively, due to the presence of 16 phenolic hydroxyls per

molecule in its structure. According to Gil et al. [77], among the po-
Extract properties

megranate ellagitannins, punicalagin, which is the largest polyphenol,

having a molecular weight of greater than 1000, is reported to be re-
sponsible for more than half the potent antioxidant activity of the juice.
143.64

Moreover, Scalbert et al. [78] reported that phenolic compounds such

138.8

as caffeic acid can easily be absorbed through the gut barrier, however
large molecular weight compounds like punicalagin and proanthocya-
nidins are very poorly absorbed. These findings are the most
Extraction yield (mg GAE/g dry peel)

convincing evidence that the antioxidant activity of pomegranate peel

is mainly related to the high contents of punicalagin in the extract.
However, there is a need for in depth study of quantitative aspect, due
to possible toxicity of this compound in large amounts of
consumption.
Recent studies showed the antioxidant properties of different po-
megranate peel extracts. Singh et al. [79] reported that a methanol
Comparison between microwave- and ultrasound-assisted extraction of phenolics from pomegranate peels. Table 2

119.82
199.4

extract of pomegranate peels showed 81% antioxidant activity at

50 ppm using the DPPH model system. In this work, the radical
Solvent/peels (mL/g) 32.2/1 35 Temperature ( C)

scavenging activity of the optimized pomegranate peel extracts for MAE

o

and UAE was 94.91 and 94.77%, respectively. The slightly lower ac-
tivity of the UAE extract may be associated with the lower punicalagin
content and the solvent used (water instead of aqueous ethanol). Negi
et al. [68], who studied the antioxidant and antimutagenic activities of
various pomegranate peel extracts, mentioned that all the extracts
showed an increase in antioxidant capacity with an increase in phe-
nolics concentration and the water extract showed less antioxidant
capacity at all the concentrations.
Power (W)Solvent/peels (mL/g)

4. Conclusions

Increased concern over the safety of synthetic antioxidants has led

to an increased interest in exploration of effective and economical
60/1

natural antioxidants. Pomegranate peel could be a good commercial

Time (min)Amplitude (W) 52

source of phenolic compounds that can be separated and concentrated

through different extraction processes. The extract can be used as a
substitute of synthetic antioxidants for food products. In this work, a
method for pomegranate peels application in food industries based on
600

the microwave-assisted extraction of phenolics compounds was opti-

Time (min)

mized. Only 4 min in aqueous ethanol (a green environmental solvent)

Ultrasound-assisted extraction
Microwave-assisted extraction

are needed to recover polyphenols from pomegranate peel with a high

Optimum extraction conditions

yield (199.4 mg GAE/g dry peel), whereas conventional extraction

10
50% aqueous ethanol 4

procedures need much longer extraction times (up to 60 times). In

addition, microwaves lead to a higher extraction yield obtained in a
shorter process time (4 min) in comparison to another “green” extrac-
tion approach, the ultrasound-assisted method (119.82 mg GAE/g dry
peel in 10 min). This difference is attributed to the intense cell de-
Solvent Water
Solvent

struction of plant material observed by SEM analysis. The obtained

extract is characterized by a great antioxidant activity (radical
Fig. 7. SEM images (×2000) of raw pomegranate peels before extraction (RM) and peels after microwave-assisted (MAE) and ultrasound-assisted (UAE) extraction at
the optimum conditions.
scavenging activity of 94.91%) due to the high content of punicalagin
(143.64 mg/g dry matter) measured by HPLC analysis. Thus, the pro-
posed method meets the terms of green process definition, since
reduces process time, allows use of alternative solvents (aqueous
ethanol) and renewable natural products, and ensure a safe and high
quality extract/ product.
Acknowledgments
This research was in part supported by the General Secretariat for
Research and Technology (GSRT) and the Hellenic Foundation for
Research and Innovation (HFRI).
References
[1] M. Çam, Y. Hişil, Pressurised water extraction of polyphenols from pomegranate
peels, Food Chem. 123 (2010) 878–885.
[2] S.R. Kanatt, R. Chander, A. Sharma, Antioxidant and antimicrobial activity of po-
megranate peel extract improves the shelf life of chicken products, Int. J. Food
Sci. Technol. 45 (2010) 216–222.
[3] B.M. Naveena, A.R. Sen, S. Vaithiyanathan, Y. Babji, N. Kondaiah, Comparative
efficacy of pomegranate juice, pomegranate rind powder extract and BHT as
anti- oxidants in cooked chicken patties, Meat Sci. 80 (2008) 1304–1308.
[4] S. Iqbal, S. Haleem, M. Akhtar, M. Zia-ul-Haq, J. Akbar, Efficiency of
pomegranate peel extracts in stabilization of sunflower oil under accelerated
conditions, Food Res. Int. 41 (2008) 194–200.
[5] O.K. Topuz, P. Yerlikaya, I. Ucak, B. Gumus, H.A. Büyükbenli, Effects of olive oil
and olive oil-pomegranate juice sauces on chemical, oxidative and sensorial quality
of marinated anchovy, Food Chem. 154 (2014) 63–70.
[6] K.V. Kumudavally, A. Tabassum, K. Radhakrishna, A.S. Bawa, Indian Pat. Appl
2009 (2009) IN 2007DE01607 A 20090306.
[7] S.K. Devatkal, P.R. Thorat, M. Manjunatha, R.K. Anurag, Comparative antioxidant
effect of aqueous extracts of curry leaves, fenugreek leaves and butylated
hydro- xytoluene in raw chicken patties, J. Food Sci. Technol. 49 (2012) 781–
785.
[8] M. Çam, F. Erdoǧan, D. Aslan, M. Dinç, Enrichment of functional properties of ice
cream with pomegranate by-products, J. Food Sci. 78 (2013) 1543–1550.
[9] J. Ventura, F. Alarcón-Aguilar, R. Roman-Ramos, E. Campos-Sepulveda, M.L.
Reyes-
Vega, V.D. Boone-Villa, E.I. Jasso-Villagómez, C.N. Aguilar, Quality and antioxidant
properties of a reduced-sugar pomegranate juice jelly with an aqueous extract of
pomegranate peels, Food Chem. 136 (2013) 109–115.
[10] H. Wasila, X. Li, L. Liu, I. Ahmad, S. Ahmad, Peel effects on phenolic composition,
antioxidant activity, and making of pomegranate juice and wine, J. Food Sci. 78
(2013) 1166–1172.
[11] K. Kaderides, A.M. Goula, K.G. Adamopoulos, A process for turning pomegranate
peels into a valuable food ingredient using ultrasound-assisted extraction and en-
capsulation, Innov. Food Sci. Emerg. Technol. 31 (2015) 204–215.
[12] H.C. Tiwari, P. Singh, P.K. Mishra, P. Srivastava, Evaluation of various techniques
for extraction of natural colorants from pomegranate rind-ultrasound and enzyme
assisted extraction, Indian J. Fibre Textile Res. 35 (2010) 272–276.
[13] Z. Pan, W. Qu, H. Ma, G.G. Atungulu, T.H. McHugh, Continuous and pulsed ul-
trasound-assisted extractions of antioxidants from pomegranate peel, Ultrason.
Sonochem. 19 (2012) 365–372.
[14] R. Tabaraki, E. Heidarizadi, A. Benvidi, Optimization of ultrasonic-assisted ex-
traction of pomegranate (Punica granatum L.) peel antioxidants by response surface
methodology, Sep. Purif. Technol. 98 (2012) 16–23.
[15] R. Boggia, F. Turrini, C. Villa, C. Lacapra, P. Zunin, B. Parodi, Green extraction from
pomegranate marcs for the production of functional foods and cosmetics,
Pharmaceuticals 9 (2016) 1–11.
[16] X. Zheng, B. Liu, L. Li, Z. Zhu, Microwave-assisted extraction and antioxidant ac-
tivity of total phenolic compounds from pomegranate peel, J. Med. Plants Res.
5 (2011) 1004–1011.
[17] J. Huang, W. He, C. Yan, X. Du, X. Shi, Microwave assisted extraction of flavonoids
from pomegranate peel and its antioxidant activity, BIO Web of Conferences 8
(2017) 1–6.
[18] P.C. Veggi, J. Martinez, M.A.A. Meireles, Fundamentals of microwave extraction, in:
F.C, G. Cravotto (Eds.), Microwave-Assisted Extraction for Bioactive Compounds:
Theory and Practice, 1st ed., Springer US, New York, 2013, pp. 15–52.
[19] X.Z. Zheng, X.W. Xu, C.H. Liu, Y. Sun, Z. Lin, H.J. Liu, Extraction characteristics
and optimal parameters of anthocyanin from blueberry powder under microwave-
as- sisted extraction conditions, Sep. Purif. Technol. 104 (2013) 17–25.
[20] K. Hayat, S. Hussain, S. Abbas, U. Farooq, B. Ding, S. Xia, C. Jia, X. Zhang, W. Xia,
Optimized microwave-assisted extraction of phenolic acids from citrus mandarin
peels and evaluation of antioxidant activity in vitro, Sep. Purif. Technol. 70 (2009)
63–70.
[21] T.S. Ballard, P. Mallikarjunan, K. Zhou, S. O’Keefe, Microwave-assisted extraction of
phenolic antioxidant compounds from peanut skins, Food Chem. 120 (2010) 1185–
1192.
[22] Y. Li, G.K. Skouroumounis, G.M. Elsey, D.K. Taylor, Microwave-assistance provides
very rapid and efficient extraction of grape seed polyphenols, Food Chem. 129
 (2011) 570–576.
[23] H. Li, Z. Deng, T. Wu, R. Liu, S. Loewen, R. Tsao, Microwave-assisted extraction of
phenolics with maximal antioxidant activities in tomatoes, Food Chem. 130 (2012)
928–936.
[24] M.D. Pavlović, A.V. Buntić, S.S. Šiler-Marinković, S.I. Dimitrijević-Branković,
Ethanol influenced fast microwave-assisted extraction for natural antioxidants ob-
taining from spent filter coffee, Sep. Purif. Technol. 118 (2013) 503–510.
[25] V.M. Simić, K.M. Rajković, S.S. Stojičević, D.T. Veličković, N.C. Nikolić, M.L. Lazić,
I.T. Karabegović, Optimization of microwave-assisted extraction of total poly-
phenolic compounds from chokeberries by response surface methodology and ar-
tificial neural network, Sep. Purif. Technol. 160 (2016) 89–97.
[26] A.R. Vieira, L. Abar, D.S.M. Chan, S. Vingeliene, E. Polemiti, C. Stevens,
D. Greenwood, T. Norat, Foods and beverages and colorectal cancer risk: a sys-
tematic review and meta-analysis of cohort studies, an update of the evidence of the
WCRF-AICR Continuous Update Project, Ann. Oncol. 28 (2017) 1788–1802.
[27] J. Pinela, M.A. Prieto, M.F. Barreiro, A.M. Carvalho, M.B.P.P. Oliveira,
J.A. Vázquez, I.C.F.R. Ferreira, Optimization of microwave-assisted extraction of
hydrophilic and lipophilic antioxidants from a surplus tomato crop by response
surface methodology, Food Bioprod. Process. 98 (2016) 283–298.
[28] S.R. Shirsath, S.S. Sable, S.G. Gaikwad, S.H. Sonawane, D.R. Saini, P.R. Gogate,
Intensification of extraction of curcumin from Curcuma amada using ultrasound
assisted approach: effect of different operating parameters, Ultrason. Sonochem.
38 (2017) 437–445.
[29] D.O. Kim, O.K. Chun, Y.J. Kim, H.Y. Moon, C.Y. Lee, Quantification of poly-
phenolics and their antioxidant capacity in fresh plums, J. Agric. Food Chem.
51 (2003) 6509–6515.
[30] C.Z. Shen, H.Y. Jun, S.H. Choi, Y.M. Kim, E.J. Jung, G.S. Oh, S.J. Joo, S.H. Kim,
I.K. Kim, Evaluation of antioxidant activities and active compounds separated from
water soluble extracts of Korean Black pine barks, Bull. Korean Chem. Soc. 31
(2010) 3567–3572.
[31] K. Kaderides, A.M. Goula, Extract from pomegranate waste as an alternative
natural antioxidant in foods. Bioprod. Process., 2019. Submitted for publication.
[32] M.M. Poojary, P. Passamonti, Extraction of lycopene from tomato processing waste:
kinetics and modelling, Food Chem. 173 (2015) 943–950.
[33] A.M. Goula, Ultrasound-assisted extraction of pomegranate seed oil - kinetic mod-
eling, J. Food Eng. 117 (2013) 492–498.
[34] J.P. Maran, V. Sivakumar, K. Thirugnanasambandham, R. Sridhar, Optimization of
microwave assisted extraction of pectin from orange peel, Carbohydr. Polym. 97
(2013) 703–709.
[35] J.H. Xie, C.J. Dong, S.P. Nie, F. Li, Z.J. Wang, M.Y. Shen, M.Y. Xie, Extraction,
chemical composition and antioxidant activity of flavonoids from Cyclocarya pa- liurus
(Batal.) Iljinskaja leaves, Food Chem. 186 (2015) 97–105.
[36] T. Wu, J. Yan, R. Liu, M.F. Marcone, H.A. Aisa, R. Tsao, Optimization of
microwave- assisted extraction of phenolics from potato and its downstream waste
using or- thogonal array design, Food Chem. 133 (2012) 1292–1298.
[37] O.R. Alara, N.H. Abdurahman, O.A. Olalere, Optimization of microwave-assisted
extraction of flavonoids and antioxidants from Vernonia amygdalina leaf using re-
sponse surface methodology, Food Bioprod. Process. 107 (2018) 36–48.
[38] C. Reichardt, Solvatochromic dyes as solvent polarity indicators, Chem. Rev. 94
(1994) 2319–2358.
[39] J.S. Boeing, É.O. Barizão, B.C. e Silva, P.F. Montanher, V. de Cinque Almeida,
J.V. Visentainer, Evaluation of solvent effect on the extraction of phenolic com-
pounds and antioxidant capacities from the berries: application of principal com-
ponent analysis, Chem. Cent. J. 8 (2014) 1–9.
[40] A.M. Galan, I. Calinescu, A. Trifan, C. Winkworth-Smith, M. Calvo-Carrascal,
C. Dodds, E. Binner, New insights into the role of selective and volumetric heating
during microwave extraction: investigation of the extraction of polyphenolic com-
pounds from sea buckthorn leaves using microwave-assisted extraction and con-
ventional solvent extraction, Chem. Eng. Process. Process. Intensif. 116 (2017) 29–
39.
[41] D.C. Campos, E.L. Dall’Oglio, P.T. De Sousa, L.G. Vasconcelos, C.A. Kuhnen,
Investigation of dielectric properties of the reaction mixture during the acid-cata-
lyzed transesterification of Brazil nut oil for biodiesel production, Fuel 117 (2014)
957–965.
[42] R. Nagahata, T. Nakamura, Microwave-assisted rapid synthesis of poly(butylene
succinate): principal effect of microwave irradiation of accelerating the poly-
condensation reaction, Polym. J. 50 (2018) 347–354.
[43] F. Anwar, R. Przybylski, Effect of solvents extraction on total phenolics and
anti- oxidant activity of extracts from flaxseed (Linum usitatissimum L.), Acta Sci.
Pol. Technol. Aliment. 11 (2012) 293–302.
[44] G. Spigno, L. Tramelli, D.M. De Faveri, Effects of extraction time, temperature and
solvent on concentration and antioxidant activity of grape marc phenolics, J. Food
Eng. 81 (2007) 200–208.
[45] F.S. Taha, G.F. Mohamed, S.H. Mohamed, S.S. Mohamed, M.M. Kamil,
Optimization of the extraction of total phenolic compounds from sunflower meal
and evaluation of the bioactivities of chosen extracts, Am. J. Food Technol. 6
(2011) 1002–1020.
[46] M. Gfrerer, E. Lankmayr, Screening, optimization and validation of microwave-
assisted extraction for the determination of persistent organochlorine pesticides,
Anal. Chim. Acta 533 (2005) 203–211.
[47] J.P. Maran, V. Sivakumar, K. Thirugnanasambandham, R. Sridhar, Microwave as-
sisted extraction of pectin from waste Citrullus lanatus fruit rinds, Carbohydr.
Polym. 101 (2014) 786–791.
[48] S.Q. Liew, G.C. Ngoh, R. Yusoff, W.H. Teoh, Sequential ultrasound-microwave as-
sisted acid extraction (UMAE) of pectin from pomelo peels, Int. J. Biol.
Macromol. 93 (2016) 426–435.
[49] J.P. Maran, K. Swathi, P. Jeevitha, J. Jayalakshmi, G. Ashvini, Microwave-
assisted extraction of pectic polysaccharide from waste mango peel, Carbohydr.
Polym. 123 (2015) 67–71.
[50]
M. Ghafoor, N.N. Misra, K. Mahadevan, B.K. Tiwari, Ultrasound assisted hydration of navy
beans (Phaseolus vulgaris), Ultrason. Sonochem. 21 (2014) 409–414.
[51] J.B. Ji, X.H. Lu, M.Q. Cai, Z.C. Xu, Improvement of leaching process of Geniposide
with ultrasound, Ultrason. Sonochem. 13 (2006) 455–462.
[52] D.M. Patil, K.G. Akamanchi, Microwave assisted process intensification and kinetic
modelling: extraction of camptothecin from Nothapodytes nimmoniana plant, Ind.
Crops Prod. 98 (2017) 60–67.
[53] R. Yedhu Krishnan, K.S. Rajan, Microwave a ssisted extraction of
flavonoids from Terminalia bellerica: study of kinetics and thermodynamics, Sep.
Purif. Technol. 157 (2016) 169–178.
[54] W. Qu, Z. Pan, H. Ma, Extraction modeling and activities of antioxidants from po-
megranate marc, J. Food Eng. 99 (2010) 16–23.
[55] F. Aqil, R. Munagala, M.V. Vadhanam, H. Kausar, J. Jeyabalan, D.J. Schultz,
R.C. Gupta, Anti-proliferative activity and protection against oxidative DNA da-
mage by punicalagin isolated from pomegranate husk, Food Res. Int. 49 (2012)
345–353.
[56] B. Gullon, M.E. Pintado, J.A. Pérez-Álvarez, M. Viuda-Martos, Assessment of
polyphenolic profile and antibacterial activity of pomegranate peel (Punica gran- atum)
flour obtained from co-product of juice extraction, Food Control 59 (2016) 94–
98.
[57] L. Yan, X. Zhou, L. Shi, D. Shalimu, C. Ma, Y. Liu, Phenolic profiles and antioxidant
activities of six Chinese pomegranate (Punica granatum L.) cultivars, Int. J. Food
Prop. 20 (2017) 94–107.
[58] T. Ismail, P. Sestili, S. Akhtar, Pomegranate peel and fruit extracts: a review
of potential anti-inflammatory and anti-infective effects, J. Ethnopharmacol.
143 (2012) 397–405.
[59] J. Lu, K. Ding, Q. Yuan, Determination of punicalagin isomers in pomegranate husk,
Chromatographia 68 (2008) 303–306.
[60] S. Kharchoufi, F. Licciardello, L. Siracusa, G. Muratore, M. Hamdi, C. Restuccia,
Antimicrobial and antioxidant features of ‘Gabsiʼ pomegranate peel extracts, Ind.
Crops Prod. 111 (2018) 345–352.
[61] H. Saad, F. Charrier-El Bouhtoury, A. Pizzi, K. Rode, B. Charrier, N. Ayed,
Characterization of pomegranate peels tannin extractives, Ind. Crops Prod. 40
(2012) 239–246.
[62] N.P. Seeram, L.S. Adams, S.M. Henning, Y. Niu, Y. Zhang, M.G. Nair, D. Heber, In
vitro antiproliferative, apoptotic and antioxidant activities of punicalagin, ellagic
acid and a total pomegranate tannin extract are enhanced in combination with
other polyphenols as found in pomegranate juice, J. Nutr. Biochem. 16 (2005)
360–367.
[63] B.H. Zhou, Z.H. Wu, X.J. Li, J. Zhang, X.M. Hu, Analysis of ellagic acid in
pome- granate rinds by capillary electrophoresis and high-performance liquid
chromato- graphy, Phytochem. Anal. 19 (2008) 86–89.
[64] P. Panichayupakarananta, A. Issuriya, A. Sirikatitham, W. Wang, Antioxidant
assay- guided purification and LC determination of ellagic acid in pomegranate
peel, J. Chromatogr. Sci. 48 (2010) 456–459.
[65] S. Rodrigues, F.A.N. Fernandes, E.S. de Brito, A.D. Sousa, N. Narain, Ultrasound
extraction of phenolics and anthocyanins from jabuticaba peel, Ind. Crops Prod. 69
(2015) 400–407.
[66] W. Elfalleh, H. Hannachi, N. Tlili, Y. Yahia, N. Nasri, A. Ferchichi, Total
phenolic contents and antioxidant activities of pomegranate peel, seed, leaf and
flower, J. Med. Plants Res. 6 (2012) 4724–4730.
[67] R. Bruni, Pomegranate phytochemicals: distribution and variability, in: A. Caligiani
(Ed.), Pomegranate Chemistry, Proceeding and Health Benefits, 1st ed., Nova
Science Publishers, USA, 2016, pp. 27–56.
[68] P.S. Negi, G.K. Jayaprakasha, B.S. Jena, Antioxidant and antimutagenic activities of
pomegranate peel extracts, Food Chem. 80 (2003) 393–397.
[69] Z. Pan, W. Qu, H. Ma, G.G. Atungulu, T.H. McHugh, Continuous and pulsed ul-
trasound-assisted extractions of antioxidants from pomegranate peel, Ultrason.
Sonochem. 18 (2011) 1249–1257.
[70] Z. Wang, Z. Pan, H. Ma, G.G. Atungulu, Extract of phenolics from pomegranate
peels, Open Food Sci. J. 5 (2011) 17–25.
[71] M. Mushtaq, B. Sultana, F. Anwar, A. Adnan, S.S.H. Rizvi, Enzyme-assisted super-
critical fluid extraction of phenolic antioxidants from pomegranate peel, J.
Supercrit. Fluids 104 (2015) 122–131.
[72] A.E. Ince, S. Sahin, G. Sumnu, Comparison of microwave and ultrasound assisted
extraction techniques for leaching of phenolic compounds from nettle, J. Food
Sci. Technol. 51 (2014) 2776–2782.
[73] M. Romdhane, C. Gourdon, Investigation in solid-liquid extraction: influence of
ultrasound, Chem. Eng. J. 87 (2002) 11–19.
[74] F. Dahmoune, B. Nayak, K. Moussi, H. Remini, K. Madani, Optimization of micro-
wave-assisted extraction of polyphenols from Myrtus communis L. Leaves, Food
Chem. 166 (2015) 585–595.
[75] Z. Lianfu, L. Zelong, Optimization and comparison of ultrasound/microwave as-
sisted extraction (UMAE) and ultrasonic assisted extraction (UAE) of lycopene from
tomatoes, Ultrason. Sonochem. 15 (2008) 731–737.
[76] M. Vinatoru, T.J. Mason, I. Calinescu, Ultrasonically assisted extraction (UAE) and
microwave assisted extraction (MAE) of functional compounds from plant mate-
rials, Trends Analyt. Chem. 97 (2017) 159–178.
[77] M.I. Gil, F.A. Tomas-Barberan, B. Hess-Pierce, D.M. Holcroft, A.A. Kader,
Antioxidant activity of pomegranate juice and its relationship with phenolic com-
position and processing, J. Agric. Food Chem. 48 (2000) 4581–4589.
[78] A. Scalbert, C. Morand, C. Manach, C. Rémésy, Absorption and metabolism of
polyphenols in the gut and impact on health, Biomed. Pharmacother. 56 (2002)
276–282.
[79] R.P. Singh, K.N.C. Murthy, G.K. Jayaprakasha, Studies on the antioxidant activity of
pomegranate (Punica granatum) peel and seed extracts using in vitro models, J.
Agric. Food Chem. 50 (2002) 81–86.