Kinetics of ultrasound-assisted extraction of antioxidant polyphenols from food by-products: Extraction and energy consumption optimization

Ultrasonics Sonochemistry 32 (2016) 137–146

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Ultrasonics Sonochemistry journal homepage: www.elsevier.com/locate/ultson

Kinetics of ultrasound-assisted extraction of antioxidant polyphenols from food by-products: Extraction and energy consumption optimization Delphine Pradal a,b, Peggy Vauchel b, Stéphane Decossin a, Pascal Dhulster b, Krasimir Dimitrov b,⇑ a b

ICAM site de Lille, 6 rue Auber, 59016 Lille Cedex, France Univ. Lille, INRA, ISA, Univ. Artois, Univ. Littoral Côte d’Opale, EA 7394 – ICV – Institut Charles Viollette, F-59000 Lille, France

a r t i c l e

i n f o

Article history: Received 16 November 2015 Received in revised form 9 February 2016 Accepted 1 March 2016 Available online 2 March 2016 Keywords: Ultrasound-assisted extraction Extraction kinetics Mathematical model Energy consumption Cichorium intybus Polyphenols

a b s t r a c t Ultrasound-assisted extraction (UAE) of antioxidant polyphenols from chicory grounds was studied in order to propose a suitable valorization of this food industry by-product. The main parameters influencing the extraction process were identified. A new mathematical model for multi-criteria optimization of UAE was proposed. This kinetic model permitted the following and the prediction of the yield of extracted polyphenols, the antioxidant activity of the obtained extracts and the energy consumption during the extraction process in wide ranges of temperature (20–60 °C), ethanol content in the solvent (0–60% (vol.) in ethanol–water mixtures) and ultrasound power (0–100 W). After experimental validation of the model, several simulations at different technological restrictions were performed to illustrate the potentiality of the model to find the optimal conditions for obtaining a given yield within minimal process duration or with minimal energy consumption. The advantage of ultrasound assistance was clearly demonstrated both for the reduction of extraction duration and for the reduction of energy consumption. Ó 2016 Elsevier B.V. All rights reserved.

1. Introduction Green extraction processes have been developed in order to overcome the problems encountered when using conventional methods. These processes are considered as environmentally friendly and promise, in particular, a shortening of processing and residence times, an acceleration of heat and mass transfers, an improvement of product quality, permit the reduction of solvent quantities and favor the use of GRAS solvents [1,2]. Therefore, the use of green extraction processes reduces energy consumption and negative impact on environment and human beings [2,3]. Ultrasound-assisted extraction (UAE) is considered as a green extraction process [3–5]. Its efficiency has been demonstrated for the extraction of bioactive compounds, providing higher recovery yields compared to classical extractions, preserving also target activities of the extracts [6,7]. In particular, UAE has been largely applied for extraction of antioxidants due to its high efficiency in terms of recovery yield and extraction rate [8–11]. Cichorium intybus L. var. sativum, belonging to Asteraceae family, is widely cultivated in Europe [12] for its roots and consumed as coffee substitute, as dried or roasted products and beverages. ⇑ Corresponding author. E-mail address: [email protected] (K. Dimitrov). http://dx.doi.org/10.1016/j.ultsonch.2016.03.001 1350-4177/Ó 2016 Elsevier B.V. All rights reserved.

Chicory roots are mainly sources of inulin and polyphenols. Inulin is a polysaccharide commonly used in different food products for fat replacement and calorie reduction [13] or as texture modifier in dairy products [14]. Polyphenols are natural antioxidants compounds [15–17] with multiple biological effects. The major property of polyphenols is their radical-scavenging capacity, which is involved in their antioxidant properties [18]. It has been reported that polyphenol-rich extracts inhibited both low-density lipoprotein (LDL) and liposome oxidation [19,20]. During processing of chicory roots several wastes are generated including chicory grounds, which are obtained after roasting of green slices, crushing into grains and extraction with hot water. Every year, about 15,000 tons of chicory grounds are generated in department of Nord (France). Nowadays, interest in reusing such wastes increases for economic and environmental considerations [21,22]. Chicory grounds could be considered as food by-products since they contain several compounds of interest, including antioxidant polyphenols, that could be valorized by a suitable extraction process. Valorization of food industry wastes as by-products is also related to the principles of the green extraction: maximum utilization of the natural resources during their processing and minimum wastes (bio-refinery concept) [2]. Similar valorizations of food byproducts by extracting antioxidant phenolics have been reported for apple pomace [6], olive wastes [22], grape marc [23] and black

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chokeberry wastes [24], for example. Most of studies on C. intybus concern inulin extraction from roots [13,25,26], few studies concern the molecular characterization of leaves and/or roots extracts [15,27–30] but, to our knowledge, none deals with extraction of phenolics from chicory by-products. Usually, to find optimal experimental conditions for extraction yield enhancement, central composite experimental design is used, taking into account parameters such as extraction time, temperature, solvent composition or power of ultrasounds [6,10,31,32]. Generally, time is included in the experimental design as a process variable, and only very few works detailed the evolution of yield as a function of time. However, it seems very important to follow the kinetics of polyphenols extraction yield and antioxidant activities in order to optimize extraction process, to decrease processing residence time and to reduce energy consumption. Recently, a mathematical model, coupling an experimental design and a description of extraction kinetics based on Peleg’s equation, has been proposed for the optimization of antioxidant polyphenols extraction from black chokeberry wastes [24]. Regarding consumed energy during the process, studies on UAE generally point out low energy consumption but only few have taken into account the real energy consumption of the process, whereas it seems important to consider it while working on the development of green processes where environmental impact is a key point. For example, JacotetNavarro et al. have compared the energy consumption at the end of extraction for some conventional and green processes [8]. To our knowledge, no work has ever been reported where energy consumption was followed during the whole extraction process, which appears necessary for optimizing the process both in terms of extraction yield and energy consumption. In the present study, the objective was to propose a green extraction process for valorization of antioxidant phenolics from a chicory by-product, and to develop a model that would enable multi-criteria optimization of the process, based on extraction yield, antioxidant activity of the extract and energy consumption during the process. After presentation of preliminary studies used to identify the main influencing parameters on extraction efficiency, the methodology to build the model for multi-criteria optimization will be developed, firstly for polyphenols yield and antioxidant activity and then for energy consumption. The interest of the proposed global model as a tool for process optimization will be demonstrated by several examples of application.

2. Materials and methods 2.1. Source – sample preparation Fresh chicory grounds were collected from a chicory-processing industrial plant based in department of Nord, in France. The moisture content of the fresh chicory grounds was 15.5 ± 0.5%. In order to obtain samples of the same moisture content for all studies and to prevent microbiological degradation, these by-products were dried for 24 h at 35 °C in an oven (Food Dehydrator, Excalibur, Lille, France). The obtained dried chicory grounds (fragments of about 0.1–0.2 cm in thickness and 0.5–1.0 cm in diameter with 92.5 ± 1.0% d.w. (dry weight)) were stored hermetically in the dark at room temperature until use. Two different batches of chicory wastes, collected in January 2014 and in April 2014, were used (lot #1 and lot #2, respectively). The total polyphenols content of each batch was characterized by decoction, at ebullition under reflux, with 2.75 g of source in 250 mL of solvent (mixture of 50% (vol.) water and 50% (vol.) ethanol). The content of polyphenols, expressed as GAE (gallic acid equivalent), was 2200 mg GAE/100 g d.w. in lot #1 and 1900 mg GAE/100 g d.w. in lot #2.

2.2. Reagents and standards As extracting solvents, deionized water or ethanol–water mixtures were used. Gallic acid (>98%), 6-hydroxy-2,5,7,8-tetramethyl chroman-2-carboxylic acid (Trolox), Folin–Ciocalteu phenol reagent (2 N), 2,20 -diphenyl-1-picrylhydrazyl (DPPH) and sodium carbonate (>99%) were supplied by Sigma–Aldrich (France), methanol (>99%) and ethanol (>99%) were provided by Flandre Chimie (France). 2.3. Extraction procedures and experimental design 2.3.1. Extraction procedures and energy consumption measurements 2.3.1.1. Preliminary studies. Preliminary studies were carried out with lot #1. The experiments on the effects of solid to liquid ratio and solvent composition were conducted in duplicate in shaking incubators for 24 h with agitation speed of 160 min1, temperature of 30 °C, and with water or ethanol–water mixtures as solvents. Solid–liquid ratio effect was studied by varying the amount of chicory grounds in 50 mL of water, with ratios of 1:10, 1:20, 1:30, 1:40 and 1:50. The influence of solvent composition was studied by varying the content of ethanol in ethanol–water mixtures from 0 to 100% (vol.) with a 10% step, at 30 °C and solid–liquid ratio of 1:30. Studies on time, temperature and ultrasonic power influences were performed in a 1 L laboratory glass extractor equipped with an ultrasound transducer at its base operating at 30.8 kHz, an agitator, a temperature regulation system and a generator of ultrasounds (SinapTec, France). The experimental setup used is depicted schematically in Fig. 1. The temperature in the extractor was maintained constant by a circulation of water in an external double-wall jacket connected to a thermostatic bath. Extraction kinetics were studied at agitation speed fixed at 160 min1 using 600 mL water as solvent and solid–liquid ratio of 1:30. When the extraction was assisted by ultrasounds, the sonication was applied in continuous mode at power of 100 W (maximal power supplied by the used ultrasonic device). The extraction kinetics were studied for a period of 2 h. Samples of the extracts were collected at 5, 10, 15, 20, 25, 30, 60, 90 and 120 min. 2.3.1.2. Optimization studies. Optimization studies were carried out with lot #2. All experiments were conducted in the laboratory ultrasound-assisted extractor (Fig. 1). The agitation speed was fixed at 160 min1, the solid–liquid ratio at 1:40, and 600 mL of water or water–ethanol mixtures were used as solvents. Experiments were carried out for a period of 2 h at three temperature levels (20, 40 and 60 °C), three levels of ethanol content in the solvent (0%, 30% and 60% (vol.) and three ultrasound power levels (0, 50 and 100 W). When ultrasound power level was studied at 100 W, the sonication was applied in continuous mode. However, when ultrasound power level was studied at the average power of 50 W, the sonication was applied in discontinuous mode (one minute on, one minute off) at power of 100 W. Energy consumption of experimental equipment during UAE were measured using an electrical meter (Otio, Auterive, France) on which were connected the ultrasound generator, the thermostatic bath and the agitator of the extractor. Samples of the obtained extracts were taken at 5, 10, 15, 20, 25, 30, 60, 90 and 120 min and the corresponding energy consumption for each sampling time was registered. 2.3.2. Experimental design A Box–Wilson procedure (Central Composite Design) was used. This methodology may identify the effects of several process variables and their interactions on several process responses, minimizing the number of experiments to be carried out. Process variables were the temperature (X1, °C), the solvent composition (X2, % ethanol content (vol.) in the solvent) and the ultrasound

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Motor

Double pale agitator

Double-wall glass extractor

Transducer

Thermostat Ultrasounds generator

Fig. 1. Schematic representation of experimental set-up used for ultrasound-assisted extraction.

Table 1 Central composite design of three variables with the observed responses and predicted by the model values for TP yield, AA of the extracts and energy consumption at the end of experimental runs (120 min of extraction). Experiment

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

Temperature, X1, °C (coded value)

Ethanol content, X2, % (vol.) (coded value)

Ultrasound power, X3, W (coded value)

TP, mg GAE/100 g

AA, lM Trolox

Observed

Predicted

Observed

Predicted

Observed

Predicted

20 20 60 60 20 20 60 60 40 40 40 40 40 20 60 40 40

0 (1) 60 (1) 0 (1) 60 (1) 0 (1) 60 (1) 0 (1) 60 (1) 30 (0) 30 (0) 30 (0) 0 (1) 60 (1) 30 (0) 30 (0) 30 (0) 30 (0)

0 (1) 0 (1) 0 (1) 0 (1) 100 (1) 100 (1) 100 (1) 100 (1) 50 (0) 50 (0) 50 (0) 50 (0) 50 (0) 50 (0) 50 (0) 0 (1) 100 (1)

655 673 826 1385 616 727 823 1532 1053 1113 1022 820 1162 724 1316 1073 1171

647 646 868 1521 685 740 871 1554 979 979 979 788 1132 694 1234 970 987

361 449 527 804 400 473 490 837 642 615 585 546 750 555 804 665 737

408 432 557 816 443 480 558 830 586 586 586 509 649 448 700 581 589

0.33 0.32 1.98 2.10 0.51 0.52 1.73 1.86 1.15 1.08 1.07 1.08 1.03 0.42 1.87 1.07 1.03

0.33 0.33 1.99 1.99 0.51 0.51 1.76 1.76 1.07 1.07 1.07 1.07 1.07 0.42 1.88 1.08 1.06

(1) (1) (1) (1) (1) (1) (1) (1) (0) (0) (0) (0) (0) (1) (1) (0) (0)

Average NRMSD

5.0%

7.2%

E, kWh

2.5%

power (X3, W). Seventeen experiments including three replicates at the center point were performed. Process responses were total polyphenols yield (TP, mg GAE/100 g d.w.), antioxidant activity of the extracts (AA, lM TEAC), and energy consumption (E, kWh). The experimental design is presented in details in Table 1. A validation of the model was performed with an experiment at conditions included in the studied experimental filed but different from the conditions used in the seventeen experiments of the experimental design, namely: X1 = +0.5, X2 = +0.5 and X3 = +0.5 (coded values).

2.4.1. Total polyphenols The concentration of total phenolic compounds in the extracts was determined using a spectrophotometer UVmini 1240 (Shimadzu France) based on Singleton et al. protocol [33]. Briefly, 0.1 mL of the liquid extracts were diluted with 7.9 mL of deionized water and mixed with 0.5 mL of Folin–Ciocalteu reagent (2 N) and 1.5 mL of 200 g L1 sodium carbonate solution. The obtained mixture was left to stand at room temperature for 2 h in the dark and the absorbance was measured at 765 nm. Results were expressed as mg GAE for 100 g d.w.

2.4. Analytical measurements

2.4.2. Antioxidant activity 2,20 -Diphenyl-1-1-picrylhydrazyl (DPPH) was used to measure the antioxidant activity of chicory grounds extracts. The method of DPPH scavenging activity employed was based on the protocol described by Brand-Williams et al. [34]. Aliquots (25 lL) of extracts were added to 975 lL of a DPPH solution in methanol (100 lM). After agitation, the reaction mixture was incubated in the dark at

For all experiments, the extracts were centrifuged for 10 min at 10,000 min1 (Eppendorf Centrifuge 5804 R, Hamburg, Germany) and the supernatants were carefully removed for further analysis. Dry matter contents were determined by drying 5 g of sample at 105 °C with a moisture balance (Precisa XM60, Poissy, France).

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room temperature for 30 min and the absorbance was measured at 517 nm by spectrophotometer UVmini 1240 (Shimadzu France). The extracts reducing activities were estimated from the decrease in absorbance. Results were expressed as lM Trolox equivalent (TEAC). 3. Results and discussion 3.1. Influence of extraction parameters on polyphenols recovery yield and antioxidant activity of the extracts While studying a solid–liquid extraction process, many parameters can have an influence on extraction efficiency, such as temperature, solvent composition [9], solid–solvent ratio [10,35], extraction time [32], solid phase particle size [36] and power of ultrasounds when applied [6,24,37]. In the present study, it was decided to use chicory grounds just as they were recovered from industry (without milling or sieving), so the particle size of the solid phase was not varied. Agitation speed was set at 160 min1 in order to obtain a homogeneous suspension of the solid particles in the solvent during extraction process. For all other parameters, preliminary studies were conducted in order to determine which ones should be taken into account in a further experimental design and to select their range of variation. 3.1.1. Effect of solid–liquid ratio It is well known that the interaction between the solvent and the solid matrix has an important influence on extraction efficiency. The volume of solvent has to be sufficient to permit a good hydration and swelling of the solid phase, leading to a better recovery yield. The effect of solid–liquid ratio on total polyphenols extraction yield from chicory grounds was studied in the range 1:10 to 1:50. The highest extraction yields were reached for ratios between 1:30 and 1:50. In the further studies on extraction of polyphenols from chicory by-products the ratios of 1:30 and 1:40 were preferred, since the use of large volumes of solvent would affect cost-efficiency of the operation. 3.1.2. Effect of solvent composition The most common solvents used for extraction from vegetal sources are water, methanol, ethanol or their mixtures [35,38]. Ethanol, which is considered as a food grade solvent, was preferred to methanol, taking into account the potential use of chicory extracts in food industry. Varying the solvent composition results in varying solvent polarity and consequently the solubility of polyphenols. Therefore, extractions with water, ethanol and water–ethanol mixtures at different ethanol content were performed in order to find optimal conditions in terms of extraction yield of total phenolics (TP) and antioxidant activity of the obtained extract (AA). Results presented in Fig 2. show very similar influence of the solvent composition on total polyphenols yield (Fig. 2a) and antioxidant activity of the obtained extracts (Fig. 2b). An increase in extraction yield and antioxidant activity was observed when ethanol content increased from 0 to 50–60%. Then, for ethanol content over 60%, the extraction yield and the antioxidant activity decreased rapidly and pure ethanol appeared to be totally inefficient. These results are in agreement with data recently reported by Galván d’Alessandro et al. [9] and Virot et al. [10], who have also investigated solubility of polyphenols in different water–ethanol mixtures and have observed highest solubility of phenolic compounds in 50% (vol.) ethanol. In the present study, best results were obtained when using 50–60% (vol.) ethanol (970 mg GAE/100 g d.w., 710 lM TEAC) and they were clearly higher than those obtained with water as solvent (720 mg GAE/100 g d.w., 480 lM TEAC).

On the base of these results, it was decided to include ethanol– water solvent composition in the studied parameters for further experimental design. 3.1.3. Effect of time, temperature and ultrasound power The influences of time, temperature and ultrasound application on extraction efficiency were studied in the same set of experiments. Kinetics of aqueous polyphenols extraction from chicory by-products were followed at two different temperatures (20 and 70 °C), without or with ultrasound assistance (0 and 100 W). Results presented in Fig. 3 show that the rate of extraction was relatively important during the first thirty minutes and then decreased progressively. As a consequence, at every studied conditions, the yield of extracted polyphenols increased with the time tending to a plateau. The extraction of phenolic compounds was highly influenced by the temperature: at 70 °C, the yields of extracted polyphenols were more than 40% higher at 30 min and about 30% higher at 120 min compared to those obtained at 20 °C. Temperature increase provides higher solubility of polyphenols, higher diffusivities in the solvent and improves mass transfer [9,39]. Similar positive effect of temperature on total polyphenols recovery during extraction from various vegetal sources has already been observed [9,39,40]. The influence of ultrasound power on extraction yield was analyzed for the two studied temperatures by comparing experiments without and with ultrasound assistance (0 and 100 W, respectively). At the two studied temperatures, a significant positive effect of the ultrasounds on the extraction rate was observed in the first thirty minutes. Then the effect of ultrasounds decreased until the end of the extraction, where it appeared very limited. While using ultrasounds, phenomenon known as acoustic cavitation occurs [41]. The cavitation force of ultrasounds can successively accelerate the heat and mass transfer rate, disrupt plants cell walls and facilitate the release of extractable compounds [3,5,42]. Ultrasounds can also facilitate swelling and hydration and cause an enlargement in the pores of the cell walls. Therefore, the diffusion process is improved and the mass transfer is enhanced [43,44]. In all preliminary studies it was observed that the influences of studied parameters (solid–solvent ratio, solvent composition, extraction time, temperature and power of ultrasounds) on antioxidant activity of the extracts were very similar to those observed for polyphenols yields. The results of these preliminary investigations showed that solvent composition, time, temperature and ultrasounds power were key parameters that had to be taken into account for further experimental design. In order to obtain higher solubility of polyphenols, it was chosen to work in the range from 0% to 60% (vol.) ethanol in the solvent. The temperature was studied between 20 and 60 °C and the ultrasound power between 0 and 100 W, respectively. 3.2. Multi-criteria optimization of UAE: model based on polyphenols extraction yield, antioxidant activity of the extract and energy consumption of the process The minimization of the process duration is a key point regarding optimization of the process. The preliminary studies showed that ultrasound assistance accelerated the extraction of antioxidant phenolics. However, the use of ultrasound assistance implies supplementary energy consumption. So, it seems essential to be able to add energy consumption as a criteria, while working on process optimization, especially in the case of development of green processes. The purpose was to propose a tool that could help finding the optimal extraction conditions on the base of TP yield, AA of the extract, and energy consumption criteria. All of those three criteria depend on operational

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Total polyphenols yield, mg GAE/100g d.w.

1200

1000

800

600

400

200

0 0

10

20

30

40

50

60

70

80

90

100

90

100

Ethanol content in the solvent, vol. %

(a) 800

Antioxidant activity, µM TEAC

700 600 500 400 300 200 100 0 0

(b)

10

20

30

40

50

60

70

80

Ethanol content in the solvent, vol. %

Fig. 2. Effect of solvent on: (a) the yield of extraction of polyphenols from chicory grounds and (b) antioxidant activity of the extract (temperature of 30 °C, solid–liquid ratio of 1:30, agitation of 160 min1, extraction time of 24 h).

conditions, but also evolve in time during the extraction. Thus, to build an efficient optimization tool it is essential to predict their kinetics (the evolution in time). Hence, the development of this optimization tool is based on modeling kinetics of those three criteria as a function of experimental conditions that were identified as relevant in the previous part of this work, namely temperature, solvent composition and power of ultrasounds. Modelling will be presented firstly for extraction kinetics for TP and AA, and then for energy consumption during the extraction. Finally, potentialities of the proposed global model as a helping tool for process optimization will be demonstrated by several examples.

3.2.1. Kinetic model for TP yields and AA of the extracts during UAE from chicory grounds The methodology chosen in the present study to model extraction kinetics in function of experimental conditions was based on the optimization model proposed by Galván d’Alessandro et al. [24], which consisted in combining Peleg’s equation and experimental design tool. The studies of extraction kinetics were based on a central composite design, composed of seventeen experiments including three central points. The whole experimental design is presented in Table 1. The extraction kinetics for TP yield and AA were described according to Eqs. (1) and (2), respectively, in which the effects of

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Total polyphenols yield, mg GAE/100g d.w.

900 800 700 600 500 400 70°C, 100 W 300 70°C, 0 W 200 20°C, 100 W 100 20°C, 0 W 0 0

20

40

60

80

100

120

140

Time, min Fig. 3. Effect of temperature and ultrasounds on the kinetics of aqueous extraction of polyphenols from chicory grounds (solid–liquid ratio of 1:30, agitation of 160 min1).

temperature, solvent composition and ultrasound power were considered in both Peleg’s coefficients K1 and K2.

TPðtÞ ¼

t 1 þ t K 1ðTPÞ K 2ðTPÞ

ð1Þ

AAðtÞ ¼

t 1 þ t K 1ðAAÞ K 2ðAAÞ

ð2Þ

1

1

where t is the extraction time (min), TP(t) is the extraction yield at time t (mg/100 g d.w.), AA(t) is the antioxidant activity (lM TEAC), K1 is Peleg’s maximal rate of extraction (at the studied conditions) and K2 is Peleg’s maximal extraction yield (at the studied conditions). All Ki coefficients (K1(TP), K2(TP), K1(AA), K2(AA)) were expressed as functions of the three studied parameters, namely temperature (X1), ethanol content in the solvent (X2) and ultrasound power (X3) and all of their interactions:

K i ¼ a0 þ a1 X 1 þ a2 X 2 þ a3 X 3 þ a11 X 21 þ a22 X 22 þ a33 X 23 þ a12 X 1 X 2 þ a13 X 1 X 3 þ a23 X 2 X 3 þ a123 X 1 X 2 X 3

ð3Þ

Experimental data obtained in these 17 experiments were used to build the model. The objective was to determine optimal values of regression coefficients (a0, a1, a2, a3, a11, a22, a33, a12, a13, a23 and a123) for each coefficient Ki, so as to minimize the deviation between experimental and model kinetic curves. To characterize this deviation it was chosen to use normalized root mean squared deviation (NRMSD) criteria:

vffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi u X u1 n 2 t ðexpp  modp Þ n NRMSD ¼

RMSD ¼ expmax

p¼1

expmax

ð4Þ

where n is the number of experimental points composing a kinetic curve (ten for each curve in the present study, corresponding to the different sampling times), expp is the experimental value at point p, modp is the model value at point p and expmax is the maximum within the n experimental values. The values of regression

coefficients for each Ki coefficient were determined using parametric estimation method with Newton algorithm, searching to minimize the mean NRMSD (the average NRMSD of the seventeen kinetics of the central composite design). Significance and suitability of the design were studied using an analysis of variance. Statistical significance of each effect was determined using Fisher test (F-value) and probability (p-value) criteria. Consequently, parameters and interactions of Eq. (3) that had no significant impact were not retained. Finally, K1(TP), K2(TP), K1(AA) and K2(AA) were calculated using Eqs. (5)–(8), respectively.

K 1ðTPÞ ¼ 100:7 þ 61:6X 1  28:1X 2 þ 10:6X 3  20:6X 1 X 2

ð5Þ

K 2ðTPÞ ¼ 1065 þ 252X 1 þ 235X 2 þ 182X 1 X 2

ð6Þ

K 1ðAAÞ ¼ 85:9 þ 56:0X 1  25:1X 2 þ 10:0X 3  24:0X 1 X 2

ð7Þ

K 2ðAAÞ ¼ 621:0 þ 109:5X 1 þ 91:6X 2 þ 67:3X 1 X 2

ð8Þ

The values of coefficients in Eqs. (5)–(8) correspond to the impacts of temperature (X1), solvent composition (X2) and ultrasound power (X3) on the maximal rate of extraction (K1) and on the maximal extraction yield (K2). The obtained values suggest that, in the studied experimental field, temperature, solvent composition and interaction between temperature and solvent composition influenced both maximal rate of extraction and maximal yield, while ultrasound power influenced maximal extraction rate but not significantly the maximal extraction yield. In Table 1 are shown the values observed and predicted by the model for TP yield and AA at the end of experimental runs (120 min of extraction) for all studied conditions. The average NRMSD (calculated on the base of all of the 17 kinetics) were about 5.0% for TP yield and 7.2% for AA, respectively, showing a good correlation between experimental and predicted kinetic curves. The good agreement between experimental and model results is illustrated in Fig. 4 for three experiments, namely the two extreme conditions (X1 = X2 = X3 = 1 and X1 = X2 = X3 = +1) and the center point of the central composite design (X1 = X2 = X3 = 0). The model was validated by performing an experiment at conditions included in the studied experimental field but different

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1800

Total polyphenols yield, mg GAE/100g d.w.

1600

60% EtOH, 60°C, 100W - exp

1400

60% EtOH, 60°C, 100W - mod

1200

Validation 45% EtOH, 50°C, 75W - exp

1000

Validation 45% EtOH, 50°C, 75W - mod

800

30% EtOH, 40°C, 50W - exp

600

30% EtOH, 40°C, 50W - mod

400

0% EtOH, 20°C, 0W - exp

200

0% EtOH, 20°C, 0W - mod

0 0

20

40

60

80

100

120

140

Time, min Fig. 4. Comparison between experimental (symbols) and simulated extraction kinetics (curves) of total polyphenols yield during UAE from chicory grounds (solid–liquid ratio of 1:40, agitation of 160 min1).

from those used in the 17 experiments of the central composite design (X1 = X2 = X3 = +0.5). Results obtained for TP yield and AA were compared to model predictions for the corresponding experimental conditions. The obtained NRMSD values of about 5.5% for TP yield and 9.5% for AA were included in the range of variation of NRMSD for the whole experimental design (2.4–10.6% for TP and 3.2–14.4% for AA, respectively). Validation results presented in Fig. 4 show the good correlation between experimental and model kinetics, proving the model ability to predict TP yield and AA of the extracts (in the studied experimental field, i.e. t = 0– 120 min, X1 = 20–60 °C, X2Validation results presented in0–60% ethanol (vol.) and X3Validation results presented in0–100 W). The comparison between Eq. (5) and Eq. (7) and between Eq. (6) and Eq. (8), respectively, shows that the influence of studied process parameters on TP yield and on AA of the extracts is very similar. This illustrates the close relationship between the content of phenolic compounds in the extracts and antioxidant activity of these extracts. 3.2.2. Kinetic model for energy consumption during UAE of antioxidant phenolics from chicory grounds To model the evolution of energy consumption during extraction process, electrical measurements were performed. For all the 17 conditions of the experimental design, it appeared that energy consumption increased linearly in function of time, so that the kinetics of energy consumption were described according to:

EðtÞ ¼ K 3 þ K 4 t

ð9Þ

where E(t) is the energy consumption (kWh) till the time t (min), K3 is the intercept (kWh) and K4 the slope (kWh min1). In fact, K3 coefficient (the intercept) corresponds to the energy consumption during the preparation phase, prior to the extraction. Indeed, the energy consumption, necessary to reach the required temperature in the solvent, was different in function of extraction conditions. The same methodology as for the modelling of TP and AA kinetics presented in the previous part was applied. Namely, K3 and K4

coefficients were expressed according to Eq. (3), to take into account the effects of the three studied variables (X1, X2, X3) and all their interactions. Values of ai coefficients were adjusted so as to minimize NRMSD (see Eq. (4)), and only variables and interactions that had a significant impact were retained (on the base of analysis of variance), leading to the expressions of K3 and K4 given in Eqs. (10) and (11):

K 3 ¼ 0:6183 þ 0:5534X 1  0:0474X 3

ð10Þ

K 4 ¼ 0:003778 þ 0:001466X 1 þ 0:000296X 3  0:000873X 1 X 3 þ 0:000642X 21

ð11Þ

The obtained values suggest that in the studied experimental field, the temperature and the ultrasound power influenced both the energy consumption during the preparation phase (K3) and the rate of energy consumption during the extraction phase (K4), while the interaction between temperature and ultrasound power and the quadratic effect of the temperature influenced K4, but not significantly K3. The solvent composition had not a significant effect on energy consumption. The good agreement between experiments and model can be observed in Fig. 5 for the same 3 examples of experiments (the two extreme conditions and the center point of the central composite design) and is assessed by the average NRMSD value of about 2.5% (calculated on the base of all 17 experiments). A validation of the proposed model for energy consumption kinetics was performed on the base of the same experiment as for TP and AA (X1 = X2 = X3 = +0.5). The measured energy consumption during validation experiment was in good agreement with the one, predicted by the model, as shown in Fig. 5 and indicated by the NRMSD value of 2.1% (included in the range of variation of NRMSD for the whole experimental design 0.4–7.6%). This confirmed the ability of the proposed model to predict energy consumption in time (in the tested experimental field).

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2 1,8

60% EtOH, 60°C, 100W - exp

1,6

Energy consumption, kWh

60% EtOH, 60°C, 100W - mod

1,4 Validation 45% EtOH, 50°C, 75W - exp

1,2 Validation 45% EtOH, 50°C, 75W - mod

1 0,8

30% EtOH, 40°C, 50W - exp

0,6

30% EtOH, 40°C, 50W - mod

0,4 0% EtOH, 20°C, 0W - exp

0,2 0% EtOH, 20°C, 0W - mod

0 0

20

40

60

80

100

120

140

Time, min Fig. 5. Comparison between experimental (symbols) and simulated (curves) energy consumption during UAE (solid–liquid ratio of 1:40, agitation of 160 min1).

Table 2 Global model results applied for minimal time and minimal energy to attain a desired yield (all simulations performed searching to reach TP = 723 mg GAE/100 g d.w., corresponding to 470 lM TEAC on average). Temperature fixed at 30 °C and ethanol content fixed at 15% (vol.)

No restriction on temperature and ethanol content

a b

Simulation conditions

Minimizing extraction duration

Minimizing energy consumption

Simulation n° Temperature (°C) Ethanol content in the solvent (%, vol.) Ultrasound power (W) Extraction duration t (min) Energy consumption at time t (kWh)

1a 60 37.5 100 9.2 1.17

3 24 60 100 59.0 0.37

2b 60 34.2 0 10.5 1.29

Minimizing extraction duration or/and energy consumption (same results) 4b 23 50.6 0 113.1 0.42

5 30 15 100 48.5 0.49

6b 30 15 0 63.5 0.55

Experimental validation of the model at these conditions. Restriction: without ultrasound assistance.

3.2.3. Global model for extraction process optimization Finally, associating Eqs. (1), (2) and (5–11), a global model is obtained, enabling to predict the evolution in time of TP yield, AA of the extract and energy consumption in function of operating conditions, i.e. temperature, solvent composition and power of ultrasounds (in the studied experimental field). The proposed global model could constitute a useful tool to optimize extraction process. For example, it can be used to predict the maximal TP yield or maximal AA of the extract that can be obtained for given operating conditions, or to find the optimal operating conditions to reach a given TP or AA for minimal extraction duration, or with minimal energy consumption. Multiple kinds of simulations with different types of constraints can be performed with this model. Some of the model potentialities will be illustrated by few simulation examples. According to the model, when carrying out the extraction in the mildest conditions (i.e. at 20 °C, with water, without ultrasounds assistance), the value of the maximal TP yield in Peleg’s equation

(K2) is about 761 mg GAE/100 g d.w., which corresponds to the maximal extractible polyphenols at these conditions if extraction would be conducted for a very long time. It was decided to fix the target TP yield at 95% of this maximal value, namely 723 mg GAE/100 g d.w. This value was chosen because it was possible to attain it at each condition of temperature between 20 and 60 °C, ethanol content in the solvent between 0% and 60% (vol.) and ultrasound power between 0 and 100 W. Hence, all examples of simulations presented below and in Table 2 were performed with the same target value for TP yield of 723 mg GAE/100 g d.w. If conducting the extraction process in the mildest conditions (i.e. at 20 °C, with water, without ultrasound assistance), more than 400 min would be necessary to attain the target TP yield. The first two simulations presented in Table 2 aimed to find the operating conditions that would enable to reach the target TP yield within the minimum process duration. Simulation results showed that the target could be attained within 9.2 min if the extraction is conducted at 60 °C, with 37.5% of ethanol in the solvent and

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ultrasound power of 100 W (simulation n°1). Hence, to obtain the target TP yield of 723 mg GAE/100 g d.w., the extraction duration could be reduced more than 40 times comparing to the extraction at the mildest conditions. A control experiment at the conditions of the simulation n°1 was carried out and the obtained values for TP yield and energy consumption after 10 min of extraction were in good agreement with model predictions (about 5% and 2% difference, respectively for TP yield and consumed energy). When the possibility of applying ultrasounds is removed (simulation n°2: US power fixed at 0 W), minimal extraction time of 10.5 min is predicted by the model (at temperature of 60 °C and 34% ethanol in the solvent). The comparison between simulations 1 and 2 shows that ultrasound assistance enables reaching the target TP yield more rapidly (more than 12% reduction of extraction time) but also with less energy consumption (about 9% reduction). The proposed global model can also be used to optimize process in term of energy consumption. Simulations 3 and 4 were performed in the view of minimizing energy consumption. To reach the target TP yield at minimal energy consumption, the model recommends to work during a little less than 1 h, at low temperature (24 °C), with 60% ethanol in the solvent and ultrasound power of 100 W (energy consumption of 0.37 kWh, simulation n°3). Very low energy consumption will be necessary also in the case without ultrasound assistance (0.42 kWh, simulation n°4), but the process duration will be almost doubled. As mentioned before, the proposed model could also permit the optimization of extraction process at various technological restrictions (no heating, aqueous extraction or reduced ethanol content in the extraction solvent, ultrasound assistance or not, etc.). For example, simulations were made for a case of relatively low temperature (30 °C) and low content of ethanol in solvent (15% vol.) with and without ultrasound assistance (simulations n°5 and 6, respectively). In this case, the optimal conditions for obtaining minimal extraction time and minimal energy consumption for the target TP yield were the same. To obtain it, 48.5 min would be necessary with the ultrasound assistance (100 W) with an energy consumption of 0.49 kWh, against 63.5 min and 0.55 kWh, respectively, in the case without ultrasounds. Hence, the use of ultrasound assistance during extraction process reduces both processing time (more than 23% in this case) and energy consumption (about 11%). The presented simulation were made on the basis of a target TP yield, but in the same manner the global model could be used to obtain minimal extraction time or minimal energy consumption necessary to obtain a target AA of the extract. Taking into account the strong relationship between TP and AA, the optimal conditions for TP and for AA are very similar. The proposed mathematical model was developed for the case of UAE of antioxidant phenolics from chicory by-products but this methodology could be easily applied for multi-criteria optimization of another kind of extraction or other mass transfer process in which it is possible to follow experimentally the kinetics of several parameters (their evolution during the process) and to describe mathematically these kinetics. According to the principles of the green extraction [2], several green impacts of the proposed UAE process could be pointed out: the use of water and agro-solvents (ethanol) only, the reduction of energy consumption during the process, the use of renewable plant sources, and the valorization of food industry wastes (byproducts).

4. Conclusion Ultrasound assisted extraction of antioxidant polyphenols is a suitable green extraction process for valorization of chicory by-

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products. The preliminary studies permitted the identification of the main parameters influencing the extraction process: time, ethanol content in solvent, temperature and ultrasound power. The very similar effect of process parameters on total polyphenols yields and on antioxidant activity of the extracts showed the strong relationship between antioxidant activity of the extracts and their content in polyphenols. The proposed new mathematical model for multi-criteria optimization permits the following and the prediction of the yield of extracted polyphenols, the antioxidant activity of the obtained extracts and the energy consumption during the extraction process for large ranges of temperature (20–60 °C), ethanol content in the solvent (0–60%) and ultrasound power (0–100 W). The advantage of ultrasound assistance was clearly demonstrated both for the reduction of extraction duration and for the reduction of energy consumption. Acknowledgement The partial financial participation of this project by Lille Métropole and Université Catholique de Lille is gratefully appreciated. References [1] C.M. Galanakis, Emerging technologies for the production of nutraceuticals from agricultural by-products: a viewpoint of opportunities and challenges, Food Bioprod. Process. 91 (2013) 575–579. [2] F. Chemat, M.A. Vian, G. Cravotto, Green extraction of natural products: concept and principles, Int. J. Mol. Sci. 13 (2012) 8615–8627. [3] N. Rombaut, A.-S. Tixier, A. Bily, F. Chemat, Green extraction processes of natural products as tools for biorefinery, Biofuels Bioprod. Biorefin. 8 (2014) 530–544. [4] T.J. Mason, L. Paniwnyk, J.P. Lorimer, The uses of ultrasound in food technology, Ultrason. Sonochem. 3 (1996) S253–S260. [5] F. Chemat, Zill-e-Huma, M.K. Khan, Applications of ultrasound in food technology: processing, preservation and extraction, Ultrason. Sonochem. 18 (2011) 813–835. [6] D. Pingret, A.-S. Fabiano-Tixier, C.L. Bourvellec, C.M.G.C. Renard, F. Chemat, Lab and pilot-scale ultrasound-assisted water extraction of polyphenols from apple pomace, J. Food Eng. 111 (2012) 73–81. [7] Z. Lianfu, L. Zelong, Optimization and comparison of ultrasound/microwave assisted extraction (UMAE) and ultrasonic assisted extraction (UAE) of lycopene from tomatoes, Ultrason. Sonochem. 15 (2008) 731–737. [8] M. Jacotet-Navarro, N. Rombaut, A.-S. Fabiano-Tixier, M. Danguien, A. Bily, F. Chemat, Ultrasound versus microwave as green processes for extraction of rosmarinic, carnosic and ursolic acids from rosemary, Ultrason. Sonochem. 27 (2015) 102–109. [9] L. Galvan d’Alessandro, K. Kriaa, I. Nikov, K. Dimitrov, Ultrasound assisted extraction of polyphenols from black chokeberry, Sep. Purif. Technol. 93 (2012) 42–47. [10] M. Virot, V. Tomao, C. Le Bourvellec, C.M.C.G. Renard, F. Chemat, Towards the industrial production of antioxidants from food processing by-products with ultrasound-assisted extraction, Ultrason. Sonochem. 17 (2010) 1066–1074. [11] M. Chen, Y. Zhao, S. Yu, Optimisation of ultrasonic-assisted extraction of phenolic compounds, antioxidants, and anthocyanins from sugar beet molasses, Food Chem. 172 (2015) 543–550. [12] H.P. Bais, G.A. Ravishankar, Cichorium intybus L. – cultivation, processing, utility, value addition and biotechnology, with an emphasis on current status and future prospects, J. Sci. Food Agric. 81 (2001) 467–484. [13] Z. Zhu, O. Bals, N. Grimi, E. Vorobiev, Pilot scale inulin extraction from chicory roots assisted by pulsed electric fields, Int. J. Food Sci. Technol. 47 (2012) 1361–1368. [14] D. Meyer, S. Bayarri, A. Tárrega, E. Costell, Inulin as texture modifier in dairy products, Food Hydrocolloids 25 (2011) 1881–1890. [15] H. Willeman, P. Hance, A. Fertin, N. Voedts, N. Duhal, J.-F. Goossens, et al., A method for the simultaneous determination of chlorogenic acid and sesquiterpene lactone content in industrial chicory root foodstuffs, Sci. World J. 2014 (2014) e583180. [16] W. Peschel, F. Sánchez-Rabaneda, W. Diekmann, A. Plescher, I. Gartzía, D. Jiménez, et al., An industrial approach in the search of natural antioxidants from vegetable and fruit wastes, Food Chem. 97 (2006) 137–150. [17] H. Liu, Q. Wang, Y. Liu, G. Chen, J. Cui, Antimicrobial and antioxidant activities of Cichorium intybus root extract using orthogonal matrix design, J. Food Sci. 78 (2013) M258–M263. [18] H. El Gharras, Polyphenols: food sources, properties and applications – a review, Int. J. Food Sci. Technol. 44 (2009) 2512–2518. [19] I.M. Heinonen, A.S. Meyer, E.N. Frankel, Antioxidant activity of berry phenolics on human low-density lipoprotein and liposome oxidation, J. Agric. Food Chem. 46 (1998) 4107–4112.

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