Continuous extraction of phenolic compounds from pomegranate peel using high voltage electrical discharge

Continuous extraction of phenolic compounds from pomegranate peel using high voltage electrical discharge

Food Chemistry 230 (2017) 354–361 Contents lists available at ScienceDirect Food Chemistry journal homepage: www.elsevier.com/locate/foodchem Analy...

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Food Chemistry 230 (2017) 354–361

Contents lists available at ScienceDirect

Food Chemistry journal homepage: www.elsevier.com/locate/foodchem

Analytical Methods

Continuous extraction of phenolic compounds from pomegranate peel using high voltage electrical discharge Jun Xi ⇑, Lang He, Liang-gong Yan School of Chemical Engineering, Sichuan University, Chengdu 610065, China

a r t i c l e

i n f o

Article history: Received 26 October 2016 Received in revised form 4 March 2017 Accepted 11 March 2017 Available online 14 March 2017 Keywords: High voltage electrical discharge Continuous extraction system Phenolic compounds Pomegranate peel Response surface methodology

a b s t r a c t Pomegranate peel, a waste generated from fruit processing industry, is a potential source of phenolic compounds that are known for their anti-oxidative properties. In this study, a continuous high voltage electrical discharge (HVED) extraction system was for the first time designed and optimized for phenolic compounds from pomegranate peel. The optimal conditions for HVED were: flow rate of materials 12 mL/ min, electrodes gap distance 3.1 mm (corresponding to 29 kV/cm of electric field intensity) and liquid to solid ratio 35 mL/g. Under these conditions, the experimental yield of phenolic compounds was 196.7 ± 6.4 mg/g, which closely agreed with the predicted value (199.83 mg/g). Compared with the warm water maceration, HVED method possessed higher efficiency for the extraction of phenolic compounds. The results demonstrated that HVED technique could be a very effective method for continuous extraction of natural compounds. Ó 2017 Elsevier Ltd. All rights reserved.

1. Introduction Pomegranate (Punica granatum L., Fig. 1a), a member of Punicaceae family, is one of the fruits containing important bioactive phenolic ingredients and has been widely used as botanical ingredients in herbal medicines and dietary supplements (Song, Li, & Li, 2016). Pomegranate fruit is rich in many nutritional and bioactive compounds, comprising organic acids, minerals (such as potassium), vitamins (C, A, and K), folic acid, etc (Akhtar, Ismail, Fraternale, & Sestili, 2015). However, the most valuable aspect of pomegranate is its phenolic compound contents, including hydrolysable tannins (punicalagins and punicalins), condensed tannins (proanthocyanidins), anthocyanins, catechins, phenolic acids (gallic, ellagic and chlorogenic), and so on (Qu, Breksa, Pan, & Ma, 2012). Phenolic compounds, which widely exist in peel, pulp and seed of pomegranate, are very beneficial to health and known for the possession of the remarkable antioxidant properties capable on protecting normal cells from various stimuli-induced oxidative stress and cell death (Nawaz, Shi, Mittal, & Kakuda, 2006). The peel (Fig. 1b) has higher content of phenolic ingredients than the pulp and seed, and can be a good source for producing high-value antioxidants (Shaban, El-Kersh, El-Rashidy, & Habashy, 2013). Pomegranate is increasingly consumed as various processed products, such as juice, wine, jam, jelly and extract, whose consumption ⇑ Corresponding author. E-mail address: [email protected] (J. Xi). http://dx.doi.org/10.1016/j.foodchem.2017.03.072 0308-8146/Ó 2017 Elsevier Ltd. All rights reserved.

has been motivated by its health benefits derived from its high antioxidant capacity (Abid et al., 2017). In pomegranate juice processing, 1 ton of fresh fruit generates 669 kg by-product pomegranate pomace containing 78% peel and 22% seed (Qu et al., 2009). Large amounts of pomegranate peel are mostly used as animal feed or discarded as useless residue, which is not only an environmental pollution but also a waste of the large raw materials (Shabtay et al., 2008). Therefore, in order to efficiently utilize pomegranate, it is necessary for us to optimize extraction of phenolic compounds from the pomegranate peel. Several studies have been published to extract phenolic compounds from pomegranate peel with various extraction methods (Kazemi, Karim, Mirhosseini, & Hamid, 2016; Masci et al., 2016; Mushtaq, Sultana, Anwar, Adnan, & Rizvi, 2015; Pan, Qu, Ma, Atungulu, & McHugh, 2012; Ranjbar, Eikani, Javanmard, & Golmohammad, 2016; Sood & Gupta, 2015; Tabaraki, Heidarizadi, & Benvidi, 2012; Çam & Hısßıl, 2010). The conventional methods are distillation and organic solvent extraction, which not only are tedious and time consuming but also exert negative impact on environment in terms of emission of organic volatile compounds (Pan et al., 2012). Also, some new and promising techniques such as pressurized water extraction (Çam & Hısßıl, 2010), instant controlled pressure drop (Ranjbar et al., 2016), supercritical fluid extraction (Mushtaq et al., 2015), and ultrasound assisted extraction (Kazemi et al., 2016) have been introduced and shown high extraction efficiency, low energy and solvent consumptions. However, little attentions have been presented on the extraction of

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Fig. 1. Pomegranate (a) and its dried peel (b).

phenolic compounds from pomegranate peel using high voltage electrical discharge (HVED). HVED is a non-thermal technique for the enhancement of mass transfer of natural ingredients in liquid at ambient temperatures and shorter extraction times by applying pulsed rapid discharge voltages (usually from 20 to 80 kV/cm electric field intensity) through an electrode gap below the surface of aqueous suspensions of natural materials, which is based on the phenomenon of electrical breakdown in liquid (Boussetta, Lebovka et al., 2009; Boussetta et al., 2011; Boussetta, Vorobiev, Reess et al., 2012; Boussetta, Turk et al., 2013; Boussetta, Lanoisellé, Bedel-Cloutour, & Vorobiev, 2009; Boussetta, Lesaint, & Vorobiev, 2013; Boussetta, Vorobiev, Le, Cordin-Falcimaigne, & Lanoisellé, 2012; Boussetta & Vorobiev, 2014; Brianceau, Turk, Vitrac, & Vorobiev, 2016; Liu, Vorobiev, Savoire, & Lanoisellé, 2011; Parniakov, Barba, Grimi, Lebovka, & Vorobiev, 2014; Rajha, Boussetta, Louka, Maroun, & Vorobiev, 2014; Sarkis et al., 2015). Electrical breakdown is the result of an avalanche of electrons turns to a starting point of streamer propagation, from the high voltage needle electrode to the grounded one, which leads to the liquid turbulence and intense mixing, the emission of high-intensity UV light, the generation of hydrogen peroxide (H2O2), the production of intense shock waves and bubble cavitation, which provoke cell structure damage and particle fragmentation, enhancing the release of intracellular components (Brianceau et al., 2016; Rajha et al., 2014). The HVED has been successfully employed for the recovery of bioactive ingredients from different plant materials including grape byproducts (Boussetta, Lebovka et al., 2009; Boussetta et al., 2011; Boussetta, Vorobiev, Reess et al., 2012; Boussetta, Lesaint et al., 2013; Boussetta, Lanoisellé et al., 2009; Boussetta, Vorobiev, Le et al., 2012; Brianceau, Turk, Vitrac, & Vorobiev, 2016; Liu, Vorobiev, Savoire, & Lanoisellé, 2011), flaxseed cake (Boussetta et al., 2013), vine shoots (Rajha et al., 2014), sesame seeds (Sarkis et al., 2015), papaya peels (Parniakov et al., 2014), and so on. These researches show that HVED can achieve less processing time, higher extraction yield, lower power consumption, less extract impurities and no harm to the activity and structure of biological active ingredient (Brianceau et al., 2016; Parniakov et al., 2014; Rajha et al., 2014; Sarkis et al., 2015). However, these investigations have been conducted using batch treatment chambers of low capacity in intermittent mode. Industries application of HVED technique requires evaluating this technique with continuous treatment chambers that allow the extraction process in continuous conditions. Response surface methodology (RSM) is a collection of statistical techniques useful for developing, improving, and optimizing complex processes, the main advantage of which was to reduce number of experimental trials needed to evaluate multiple variables and their interactions (Eren & Kaymak-Ertekin, 2007). Thus, the objective of this work is to develop a continuous

HVED system for extraction of phenolic compounds from pomegranate peel and to optimize the extraction process using RSM. 2. Materials and methods 2.1. Plant materials and reagents Pomegranate fruits were purchased from local markets in Chengdu, China. Pomegranate fruits were washed with distilled water, and then manually peeled and their edible portions were carefully separated. The peels were dried in a hot air oven at 40 °C for 48 h, grounded to a fine powder using a grinder machine (KC-1000, Beijing Kaichuangtonghe Technology Development Co., Ltd, Beijing, China) and passed through a 100 mesh sieve, then packaged in polyethylene bags and store in refrigerator at 4 °C until used. Ethanol used in the experimental work was analytical reagent grade chemicals (Beijing Chemical Reagents Company, Beijing, China). DPPH free radical was purchased from Sigma Chemical Co. (St Louis, USA). Gallic acid, Folin-Ciocalteu’s phenol reagent, and Na2CO3 were purchased from the Sinopharm Chemical Reagent Co. (Beijing, China). Other reagents were of analytical grade and purchased from Chengdu Chemical Industry (Chengdu, China). All solutions were prepared with analytical chemicals and deionized water was used throughout. The UV–Vis spectrophotometer (751-GW) was from Shanghai Analytical Instrument Overall Factory (Shanghai, China). 2.2. Design of the continuous HVED extraction system The HVED extraction system (Fig. 2a) was composed of a highvoltage pulse generator, a continuous treatment chamber (Fig. 2b), a data acquisition systems, a fluid handling and cooling system, and voltage and current measuring devices. The high-voltage pulse generator (TP3020) was obtained from Dalian Teslaman Technology Co., Ltd. (Dalian, China). This pulse generator offered exponentially decaying bipolar triangle waveform pulses with 2 ls pulse duration and frequency up to 1000 Hz, which provided 20 kV–10 kA discharges for a few microseconds. The ‘‘converged electric field type” treatment chamber described by Alkhafaji and Farid was used (Fig. 2b), which provided higher electric field intensity in a small volume without increasing the voltage at the electrodes (Alkhafaji & Farid, 2007). The chamber contained a pair of parallel disc mesh electrodes made of stainless steel and an insulating plate with a small hole between the electrodes to create a small orifice (i.e., treatment region) where electric field intensity was highly concentrated. The electric field

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Fig. 2. Schematic representation of a continuous HVED extraction system (a) and Photograph of UV light appearing in the treatment region at 20 kV/cm electric field intensity (b (1)) and cross-sectional view of the treatment chamber (b (2)).

intensity in the center hole of the insulating plate was much larger than other parts of the chamber. Therefore the insulator center hole was a region of HVED processing center. The stainless steel electrode received high voltage pulse power at one end, and was grounded at the other end. The liquid materials were continuously pumped and introduced into the chamber through the holes of the disc electrodes, and only the materials inside the hole of the insulating plate were subjected to the HVED treatment. A photograph of UV light appearing in the treatment region at 20 kV/cm electric field intensity was shown in Fig. 2b (1). The diameters of both electrodes and the treatment region were 20 mm and 1 mm, respectively. The diameter of the mesh opening was 2 mm. The distance between the electrodes d varied from 1 to 10 mm. The values of 2, 3, 4 and 5 mm were tested, and the 9 kV peak pulse voltage was used. The electric field intensity (E, kV/cm) was calculated using the following equation:



V d

ð1Þ

where, v: peak pulse voltage (kV), d: electrodes gap distance (cm). 2.3. Continuous HVED extraction process Many researches had demonstrated that water was an environmentally friendly and efficient solvent for extracting phenolic

compounds from plant materials (Pan et al., 2012; Ranjbar et al., 2016). Therefore, the water was used as the extraction solvent in this research. In order to operate HVED efficiently, flow rate of materials (8–14 mL/min), electrodes gap distance (2–5 mm, these distances correspond with electric field intensity in the range of 18–45 kV/cm) and liquid to solid ratio (20–50 mL/g) have to be carefully controlled for a continuous HVED system. 10 g of the dried pomegranate peel samples with appropriate volume of distilled water were finely blended using a stirrer (JJ-1, Hongke Instrument Factory, Jintan, China). The suspension was pumped into a cooling coil immersed in water bath before entering the HVED treatment chamber using a peristaltic pump (WG600S, Baoding Lead Fluid Technology Co., Ltd, Baoding, China) with appropriate flow rate. Then, the suspension was subjected to electric discharge treatment at different distances between the electrodes with a constant peak pulse voltage of 9 kV and frequency of 100 Hz. The treated suspension leaving the chamber and passed through the cooling coil were collected immediately. After extraction, the mixture was filtered through the quantitative filter paper (8–10 mm, Whatman, UK). Then, the extracting solution was centrifuged at 4000 rpm for 10 min and the supernatant was carefully gathered by membrane filtration (0.45 lm, Millipore, USA). The filtrate was collected and stored at 4 °C in refrigerator for subsequent analysis. Warm water maceration was performed as a control experiment. According to the preliminary optimized investigation (data

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not shown), the optimum condition of warm water maceration was as follows: temperature 70 °C, time 60 min, liquid to solid ratio 35 mL/g.

3. Results and discussion

2.4. Determination of the yield of phenolic compounds The yield of phenolic compounds was determined using FolinCiocalteu’s method (Li et al., 2006). 1 mL of the extract was mixed with 4 mL of 7.5% Na2CO3 and 5 mL of 10-fold diluted FolinCiocalteu reagent. After 60 min at 25 °C, the absorbance was measured at 765 nm by a UV–Vis spectrophotometer (751-GW). The blank had all reagents, but the extract was replaced by distilled water. Gallic acid was used as standard to plot a calibration curve. All analyses were carried out in triplicate and the results were expressed in mg gallic acid equivalent/g dried sample.

2.5. Experimental design In order to identify the independent variables and appropriate ranges of the Box-Behnken design, some single-factor experiments were carried out and the effects of each factor on the extraction were analyzed and evaluated by determining the yield of phenolic compounds. According to the obtained results of above experiments, we adopt a three-variable, three-level Box-Behnken design with response surface methodology to optimize the extraction of phenolic compounds. The three independent variables was flow rate of materials (mL/min, x1), electrodes gap distance (mm, x2), and liquid to solid ratio (mL/g, x3). The yield of phenolic compounds (mg/ g, y) was considered be the response of the design experiments. Each variable set at the three levels. Seventeen experiments were carried out with three times at the center points to evaluate the pure error. The design plan was showed in Table 1. The experimental data was fitted to the following second-order polynomial model:

y ¼ A0 þ

3 3 2 X 3 X X X A i xi þ Aii x2i þ Aij xi xj i¼1

i¼1

The experimental design, graphical and statistical analysis were performed using Design-Expert 8.0.6 (Trial version, Stat-Ease Inc., Minneapolis, USA). All trials were performed in triplicate.

ð2Þ

i¼1 j¼iþ1

where, y was the estimated response; xi and xj were the independent variable influencing the responses function y; A0, Ai, Aii and Aij represented the regression coefficients for intercept, linear, quadratic and interaction terms, respectively.

3.1. Single-factor experiments In this study, the flow rate of materials, distance between the electrodes and liquid to solid ratio were studied, respectively. Fig. 3 showed the effects of these factors on the yield of phenolic compounds. 3.1.1. Effect of flow rate of materials In order to determine the optimum flow rate of materials, 10 g of the dried material sample was extracted with 300 mL of distilled water. The electrodes gap distance was fixed to 3 mm (30 kV/cm of electric field intensity). The flow rate of materials was 8, 10, 12 and 14 mL/min (corresponding to 39, 31, 26, 22 min of extraction duration), respectively. Fig. 3a showed the effect of flow rate of materials on the yield of phenolic compounds from pomegranate peel. It can be seen that the phenolic compounds yield increased as the flow rate of materials was increased from 8 to 10 mL/min, because the thickness of the boundary layer decreased with the increase of flow rate and more phenolic compounds was extracted into the solvent (Psillakis & Kalogerakis, 2001). Further increases in flow rate of materials resulted in decrease in the yield of phenolic compounds. This is possibly due to the linear velocity of the sample solution being too high to reach the extraction equilibrium in the boundary layer of both phases (Liu, Chen, Yang, & Wang, 2007). Therefore, the ranges of 8–12 mL/min were used for further optimization. 3.1.2. Effect of distance between the electrodes To study the effects of the electrodes gap distance on the yield of phenolic compounds, different electrodes gap distances of 2, 3, 4 and 5 mm (corresponding to 45, 30, 22.5, and 18 kV/cm of electric field intensity calculated from the Eq. (1)) were used, accompanied by a liquid to solid ratio of 30 mL/g, and a flow rate of materials of 10 mL/min (corresponding to 31 min of extraction duration). The electric field intensity was adjusted by changing distance between the electrodes. The wider the distance was, the lower the electric field intensity became. For discharges formation, a small electrodes gap distance is necessary because of the relatively high breakdown electric field of water (Brianceau et al., 2016; Liu et al., 2011).

Table 1 The Box-Behnken design for optimizing extraction conditions. Run

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17

Independent variable

y: The yield of phenolic compounds (mg/g)

x1: Flow rate of materials (mL/min)

x2: Distance between the electrodes (mm)

x3: Liquid to solid ratio (mL/g)

10 8 12 10 10 8 10 10 12 12 10 10 8 12 10 10 8

4 2 3 3 3 3 3 3 4 2 2 2 4 3 4 3 3

20 30 40 30 30 20 30 30 30 30 20 40 30 20 40 30 40

113.57 105.31 199.42 179.60 181.02 117.30 178.35 183.42 172.87 128.25 90.46 114.11 139.70 100.25 103.45 180.22 95.78

The yield of phenolic compounds (mg/g)

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180

(a)

160 140 120 100 80 60 40 20 0

The yield of phenolic compounds (mg/g)

8

180

14

3 4 Distance between the electrodes (mm)

5

(b)

160 140 120 100 80 60 40 20 0 2

The yield of phenolic compounds (mg/g)

10 12 Flow rate of materials (mL/min)

180

(c)

160 140 120 100 80 60 40 20 0 20

30 40 Liquid to solid ratio (mL/g)

50

Fig. 3. The effect of different flow rate of materials (a), distance between the electrodes (b), and liquid to solid ratio (c) on the yield of phenolic compounds. The peak pulse voltage was fixed to 9 kV. Data represent means ± SD of three independent experiments.

Fig. 3b showed that the yield of phenolic compounds increases when the electrodes gap distance increased from 2 to 3 mm, and then decreases when the electrodes gap distance was more than 3 mm, that is, when the electric field intensity increased from 18 to 30 kV/cm, the yield of phenolic compounds rose steadily, and then dropped from 30 to 45 kV/cm. At a small electrodes gap distance of 3 mm, the arc discharge occurred at a very short time. If the gap is more than 3 mm, the discharge intensity would be reduced because of the weakened electric field. Therefore, the electrodes gap distance of 3 mm was considered to be optimal in the present experiment. The experimental result indicated that the electric field intensity had a strong effect on the yield of phenolic compounds. These results were in accordance with previous studies conducted in batch system (Brianceau et al., 2016; Rajha et al., 2014; Sarkis et al., 2015). The HVED enhancement of the extraction process might be due to the augment of electrical breakdown potential of the cell membrane. The higher electric field intensity could cause the more serious damages of the cell. Thus, it is easier to transfer solute from the cell to water (Boussetta, Lesaint et al., 2013; Boussetta & Vorobiev, 2014; Brianceau et al., 2016). However, the yield of phenolic compounds decreased rapidly when the electric field intensity was greater than 30 kV/cm. This was probably because the higher electric field intensity might cause the severe disruptions of structure of phenolic compounds, which we will further investigate in the future. 3.1.3. Effect of liquid to solid ratio The effect of the liquid to solid ratio on the extraction process was investigated at 20, 30, 40 and 50 mL/g, respectively. At the same time, the flow rate of materials was set to 10 mL/min (corresponding to 31 min of extraction duration) and distance between the electrodes was set to 3 mm (30 kV/cm of electric field intensity). It could be found from Fig. 3c that the yield of phenolic compounds went up steadily with the increase of the liquid to solid ratio from 20 to 30 mL/g until the critical value at 30 mL/g, and then decreased mildly above 30 mL/g. A possible explanation is that a larger volume of solvent dissolves a larger quantity of phenolic compounds, which results in a higher extraction yield of the phenolic compounds (Zhang, Yang, & Wang, 2011). But if the liquid to solid ratio reached considerable extend, the excess of solvent could absorb energy from the extraction system and thereby lower the yield of phenolic compounds. Therefore, in order to achieve more phenolic compounds with less solvent, 30 mL/g of liquid to solid ratio was chosen in the present experiment.

Table 2 ANOVA for the response surface quadratic model. Source

Sum of square

Degree of freedom

Mean square

F-value

p-Value

Significance

Model x1 x2 x3 x21 x22 x23 x1x2 x1x3 x2x3 Lack-of-fit Pure error R2 Adj. R2

233.53 18.74 0.38 0.44 4.75 46.87 73.25 0.26 36.42 2.82 118.83 0.14 0.9994 0.9976

12 1 1 1 1 1 1 1 1 1 3 4

19.46 18.74 0.38 0.44 4.75 46.87 73.25 0.26 36.42 2.82 394.61 0.036

544.16 524.13 10.56 12.42 132.90 1310.62 2048.21 7.32 1018.23 78.97 11.03

<0.0001 <0.0001 0.0314 0.0244 0.0003 < 0.0001 < 0.0001 0.0538 < 0.0001 0.0009 0.1038

**

x1: Flow rate of materials (mL/min), x2: Distance between the electrodes (mm), x3: Liquid to solid ratio (mL/g). * Significant coefficient (p < 0.05). ** Highly significant coefficient (p < 0.01).

** * * * ** ** * ** **

Not significant

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Fig. 4. Response surface plots for the effect of the distance between the electrodes and flow rate of materials (a), liquid to solid ratio and flow rate of materials (b), liquid to solid ratio and distance between the electrodes (c) on the yield of phenolic compounds. The peak pulse voltage was fixed to 9 kV.

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3.2. Optimization of the extraction progress 3.2.1. Analysis of the model According to the method of Box-Behnken design and the singlefactor experiments, the flow rate of materials (mL/min, x1), distance between the electrodes (mm, x2), and liquid to solid ratio (mL/g, x3) which have a great effect on the yield of phenolic compounds were selected as design variables in the RSM. The values of yields of phenolic compounds at different experimental combinations were given in Table 1. After analysis, the regression model could be described by the following quadratic polynomial in terms of actual values.

y ¼ 503:3 þ 12:82x1 þ 224:31x2 þ 13:62x3 þ 1:27x1 x2 þ 1:51x1 x3  0:84x2 x3  2:65x21  33:39x22  0:42x23

ð3Þ

where, y was the yield of phenolic compounds (mg/g), x1 was the flow rate of materials (mL/min), x2 was the distance between the electrodes (mm) and x3 was the liquid to solid ratio (mL/g). It was required to test the significance and adequacy of the model through performing in the form of analysis of ANOVA for the quadratic model. Table 2 showed that the model of p-value (p < 0.0001), correlation coefficient (R2 = 0.9994), and adjusted correlation coefficient (Adj. R2 = 0.9976), which means that it was a high precision and applicable model (Boussetta & Vorobiev, 2014; Sarkis et al., 2015). From the p-values of each model term (all lower than 0.05), it could be concluded that the linear coefficients (x1, x2 and x3), interactive coefficient (x1x2, x1x3 and x2x3) and quadratic coefficients (x21, x22 and x23) were significant for phenolic compounds extraction (Boussetta, Lesaint et al., 2013; Brianceau et al., 2016). 3.2.2. Analysis of response surfaces The relationship between independent and dependent variables was illustrated in 3D plots of the response surfaces generated by the model (Fig. 4). Two variables were depicted in one 3D surface plot while the other variable was kept at its central level. It was very easy to understand the interactions between two variables and also to determine their optimum levels by these 3D plots (Brianceau et al., 2016). Fig. 4a showed the 3D plot for different flow rate of materials and distance between the electrodes when the liquid to solid ratio was fixed at 30 mL/g. It could be seen that the yield increased at first as the flow rate of materials increased from 8 to 10 mL/min or the distance between the electrodes increased from 2 to 3 mm, but it decreased when these two variables kept increasing thereafter. The interaction of liquid to solid ratio and flow rate of materials on phenolic compounds extraction was shown in Fig. 4b, for a fixed distance between the electrodes of 3 mm. The yield increased quickly to its maximum as both liquid to solid ratio and flow rate of materials initially increased, and it decreased thereafter. Fig. 4c showed the relationship between the phenolic compounds yield and the two variables of liquid to solid ratio and distance between the electrodes for a fixed flow rate of materials of 10 mL/min. When the flow rate of materials was set, the yield improved significantly as the liquid to solid ratio increased from 20 to 30 mL/g and then began to decrease slowly. Meanwhile, the yield increased rapidly with increasing distance between the electrodes from 2 to 3 mm, then decreased quickly as the distance between the electrodes increased further. 3.2.3. Optimal conditions and verification of predictive model According to Fig. 4, the tested variables for obtaining the maximum yield of phenolic compounds were calculated by the software Design Expert, and the derived optimum values were as

follows: flow rate of materials 12 mL/min (26 min of extraction duration), distance between the electrodes 3.15 mm and liquid to solid ratio 34.83 mL/g. Under these optimal conditions, the maximum predicted yield of phenolic compounds was 199.83 mg/g. For simplicity, the flow rate of materials 12 mL/min (26 min of extraction duration), distance between the electrodes 3.1 mm (29 kV/cm of electric field intensity) and liquid to solid ratio 35 mL/g were used as the optimal conditions. Under these conditions, the experimental yield of phenolic compounds was 196.7 ± 6.4 mg/g, which agreed closely with the predicted value. The good correlation between experimental and predicted values confirmed that the response model was accurate and adequate for the extraction of phenolic compounds (Boussetta, Lesaint et al., 2013; Brianceau et al., 2016; Parniakov et al., 2014; Rajha et al., 2014). 3.3. Comparison between continuous HVED and warm water maceration The extraction efficiency of the phenolic compounds was compared by continuous HVED and warm water maceration under the optimal conditions. The results indicated that the yield of phenolic compounds (196.7 ± 6.4 mg/g) by continuous HVED for 30 min was significantly higher than that (158.9 ± 7.2 mg/g) by warm water maceration for 60 min. That is to say, there was 23.78% increase in the yield as a result of using continuous HVED procedure. Therefore, the findings confirmed the extraction enhancing effect of HVED treatment. 4. Conclusions In this study, a continuous HVED extraction system of phenolic compounds from pomegranate peel was developed and optimized, and a comparison with conventional extraction method was carried out. HVED technique gave a significant increase in the yield, which seemingly ascribed to enhanced mass transfer resulted from disruptions of sample cells under electrical breakdown. Thus, the results indicated that HVED extraction method was a promising technique for extracting phenolic compounds from pomegranate peel. Acknowledgement This work is financially supported by the National Natural Science Foundation of China (No. 21376150), Postdoctoral Science Foundation of China (No. 2013M530400, 2014T70871) and the Specialized Research Fund for the Doctoral Program of Higher Education of China (No. 20100181120076). References Abid, M., Cheikhrouhou, S., Renard Catherine, M. G. C., Bureau, S., Cuvelier, G., Attia, H., & Ayadi, M. A. (2017). Characterization of pectins extracted from pomegranate peel and their gelling properties. Food Chemistry, 215, 318–325. Akhtar, S., Ismail, T., Fraternale, D., & Sestili, P. (2015). Pomegranate peel and peel extracts: Chemistry and food features. Food Chemistry, 174, 417–425. Alkhafaji, S. R., & Farid, M. (2007). An investigation on pulsed electric fields technology using new treatment chamber design. Innovative Food Science and Emerging Technologies, 8(2), 205–212. Boussetta, N., Lanoisellé, J. L., Bedel-Cloutour, C., & Vorobiev, E. (2009). Extraction of soluble matter from grape pomace by high voltage electrical discharges for polyphenol recovery: Effect of sulphur dioxide and thermal treatments. Journal of Food Engineering, 95, 192–198. Boussetta, N., Lebovka, N., Vorobiev, E., Adenier, H., Bedel-Cloutour, C., & Lanoisellé, J. L. (2009). Electrically assisted extraction of soluble matter from chardonnay grape skins for polyphenol recovery. Journal of Agricultural and Food Chemistry, 57, 1491–1497. Boussetta, N., Lesaint, O., & Vorobiev, E. (2013). A study of mechanisms involved during the extraction of polyphenols from grape seeds by pulsed electrical discharges. Innovative Food Science and Emerging Technologies, 19, 124–132.

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