Application of Ultrasound in Grape Mash Treatment in Juice Processing

Ultrasonics Sonochemistry xxx (2009) xxx–xxx

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Ultrasonics Sonochemistry

journal homepage: www.elsevier .com/ locate /ul tsonch

Application of ultrasound in grape mash treatment in juice processing

Le Ngoc Lieu, Van Viet Man Le *

Dep. of Food Tech., Ho Chi Minh City University of Technology, Ho Chi Minh City, Viet Nam

a r t i c l e i n f o

Article history:Received 15 January 2009Received in revised form 25 April 2009Accepted 7 May 2009Available online xxxx

PACS:43.35.+d47.35.Rs62.60.+v81.40.Gh83.80.Mc83.85.Jn

Keywords:Enzymatic treatmentGrape mashOptimizationUltrasound

1350-4177/$ - see front matter � 2009 Elsevier B.V. Adoi:10.1016/j.ultsonch.2009.05.002

* Corresponding author. Tel.: +84 8 38 64 62 51; faE-mail address: [email protected] (V.V.M. Le)

Please cite this article in press as: L.N. Lieu, V.V.doi:10.1016/j.ultsonch.2009.05.002

a b s t r a c t

Recently, application of ultrasound has attracted considerable interest as an alternative approach to tra-ditional methods. In this study, response surface methodology (RSM) was used to optimize the conditionsfor grape mash treatment by ultrasound and by combination of ultrasound and enzyme. The results indi-cated that optimal conditions were the temperature of 74 �C and the time of 13 min for sonication treat-ment; and were the enzyme concentration of 0.05% and the time of 10 min for combined ultrasound andenzyme treatment. In comparison with traditionally enzymatic treatment, sonication treatmentincreased extraction yield 3.4% and shortened treatment time over three times; combined ultrasoundand enzyme treatment increased extraction yield slightly, only 2%, but shortened treatment time overfour times. After sonication treatment, enzymatic treatment increased extraction yield 7.3% and totaltreatment time of this method was still shorter than that of traditionally enzymatic treatment method.Besides, application of ultrasound improved the grape juice quality because it increased contents of sug-ars, total acids and phenolics as well as color density of grape juice.

� 2009 Elsevier B.V. All rights reserved.

1. Introduction

Grape juice is not consumed in large amounts because it is toosweet or too acidic [1]. However, grape is the single most abundantfruit harvested in the world [2] because grape wines are producedin greatest volume [1]. Traditionally, grape mash is treated withenzymes to increase volume of free-run juice and to reduce press-ing time. However, enzymatic maceration takes much time [3] andtherefore the cost of energy is increased.

Recently, application of ultrasonic technology in food process-ing has widely attracted attentions. Ultrasound was applied inextraction of plant materials because of enhancement of yieldand shortening of extraction time [4–6]. There are several studieson application of ultrasound in extraction, but the authors wereinterested in one or two valuable components in the plant extractsuch as phenolics [7–9], tartaric and malic acids [10], flavors [11–13], lycopene [14], oil [15,16], polysaccharides [17–20]. None ofthese studies mentioned simultaneous extraction of many com-pounds by ultrasound in juice processing. In addition, ultrasoundwas applied in enzymatic treatment because of its ability of violentagitation and its positive effects on enzyme activity [21–26].

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M. Le, Application of ultrasound

However, there are no studies on application of ultrasound in enzy-matic treatment of fruit mash in juice processing.

The objective of this study was to determine optimal conditionsof ultrasound assisted process and combined ultrasound and en-zyme process for grape mash treatment by using response surfacemethodology as well as to compare efficiency of these treatmentmethods with that of traditionally enzymatic method.

2. Materials and methods

2.1. Materials

2.1.1. Enzyme sourcePectinex Ultra SP-L from Aspergillus aculeatus obtained from

Novozymes Switzerland AG, Dittengen, Switzerland – was usedin this study. This enzyme preparation contains different pectino-lytic enzymes [endo-polygalacturonase (EC 3.2.1.15; C.A.S. No.9032-75-1), pectin-lyase (EC 4.2.2.10; C.A.S. No. 9033-35-6),pectin esterase (EC 3.1.1.11; C.A.S. No. 9025-98-3)], and otheractivities, such as b-galactosidase, cellulase, chitinase and trans-galactosidase [27]. The activity of Pectinex Ultra SP-L is 26,000PG per mL (polygalacturonase activity per mL). The catalytic tem-perature and pH of this enzyme preparation are 50 �C and 4.5,respectively [28–30].

in grape mash treatment in juice processing, Ultrason. Sonochem. (2009),

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Table 2Experimental planning and results of extraction yield for sonication treatment ofgrape mash.

Run Temperature (�C) Time (min) Yield (%)

1 60 5 74.92 80 5 80.33 60 15 79.34 80 15 81.05 55.9 10 75.86 84.1 10 80.47 70 2.9 75.98 70 17.1 82.29 70 10 81.510 70 10 81.411 70 10 81.212 70 10 81.813 70 10 81.3

2 L.N. Lieu, V.V.M. Le / Ultrasonics Sonochemistry xxx (2009) xxx–xxx

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2.1.2. Grape mashGrape (Red Cardinal) used in this study was purchased from a lo-

cal market in Ninh Thuan, Vietnam. Grape was destemmed,washed and crushing in a blender (National, Vietnam) for 2–3 min. Then the pH of grape mash was adjusted to value of 4.5.

2.2. Experimental methods

2.2.1. Enzymatic treatmentSamples of 250 mL grape mash were taken for each assay. The

samples were placed into 500 mL flasks.First series: Different amounts of Pectinex Ultra SP-L were

added into flasks of samples. Enzyme concentration was variedfrom 0%v/v to 0.1%v/v. The samples were then kept in the periodof 40 min.

Second series: Pectinex Ultra SP-L (0.04%v/v) was added intoflasks of samples. The treatment time was varied from 10 to60 min.

In both series, treatment temperature was adjusted to 50 �C byusing a thermostatic water bath (Memmert, WNB 45, Yogyakarta,Indonesia). At the end of the process, enzymes in the sample wereinactivated by heating the mash at 90 �C for 5 min in a water bath.The mash was then filtered through a cheese cloth. The obtainedsuspension was centrifuged at 6500 rpm for 10 min by a refriger-ated centrifuge (Sartorius, Sigma 3K30, Geneva, Switzerland) andthe supernatant was collected for further analysis.

2.2.2. Sonication treatmentA randomised, quadratic central composite circumscribed (CCC)

response surface design was used to study the effect of tempera-ture and treatment time on the extraction yield of grape mashtreatment by ultrasound. The software Modde version 5.0 wasused to generate the experimental planning and to process data.

For each assay, 2 L grape mash with total solid content ofapproximately 20% was directly poured into an ultrasonic bath.The height of the mash in the bath was about 4.5 cm. The bath(Elma�, T 660/H, Singen, Germany) is a rectangular container(300 � 151 � 150 mm) with the maximal volume of 5.75 L, towhich 35 kHz transducers are annealed at the bottom so that ultra-sonic waves are transmitted from the bottom to above. The equip-ment operated at an ultrasound intensity of 2 W/cm2 and anultrasound power of 360 W. The sonotrode of the bath had a sur-face area of about 180 cm2 which was large enough for ultrasonicwave to distribute homogeneously in the height of the treatedsample. The bath was equipped with a thermostatic system.

The treatment temperature was ranged from 60 to 80 �C and thetime was ranged from 5 to 15 min. The experimental design is pre-sented in Table 2. At the end of the process, the mash was also fil-tered and centrifuged in the same way of Section 2.2.1.

2.2.3. Combined ultrasound and enzyme treatmentIn this treatment, grape mash was simultaneously treated by

ultrasound and enzyme in the ultrasonic bath. A randomised, qua-dratic central composite circumscribed (CCC) response surface de-sign was also used to study the effect of enzyme concentration and

Table 1Independent variables and their levels in the response surface design.

Process Independent variables

Ultrasound assisted treatment Temperature (�C)Time (min)

Combined ultrasound and enzyme treatment Enzyme concentration (%v/v)Time (min)

Please cite this article in press as: L.N. Lieu, V.V.M. Le, Application of ultrasounddoi:10.1016/j.ultsonch.2009.05.002

treatment time on the extraction yield. The software Modde ver-sion 5.0 was also used to generate the experimental planningand to process data.

For each assay, 2 L grape mash was added directly into theultrasonic bath. A determined amount of Pectinex SP-L (from0.02%v/v to 0.06%v/v) was added and the mixture was stirred be-fore treatment. The treatment time was ranged from 4 to 12 min.The experimental design is presented in Table 5. Temperaturewas maintained at 50 �C. At the end of the treatment, enzymes inthe sample were inactivated by heating the mash at 90 �C for5 min in a water bath. The following steps were similar to thosein Section 2.2.1.

2.2.4. Enzymatic treatment after sonicationThe samples obtained from the experiments of ultrasonic treat-

ment (Section 2.2.2) were then treated with Pectinex Ultra SP-L.This part consisted of two series of experiments. For each assay,samples of 250 mL grape mash were taken and placed into500 mL flasks.

First series: different amounts of Pectinex Ultra SP-L wereadded into flasks of samples. Enzyme concentration was variedfrom 0%v/v to 0.1%v/v. The samples were then kept in the periodof 20 min.

Second series: Pectinex Ultra SP-L (0.06%v/v) was added intoflasks of samples. The treatment time was ranged from 10 to40 min.

In both series, temperature was maintained at 50 �C. The fol-lowing steps were similar to those in Section 2.2.1.

2.2.5. Comparison in physico-chemical characteristics of grape juiceobtained from different grape mash treatment methods

In order to compare some physico-chemical characteristics ofgrape juice obtained from different grape mash treatment meth-ods, all experiments were carried out again at the appropriate con-ditions obtained from Section 2.2.1 to 2.2.4. The obtained sampleswere further analyzed in reducing sugar content, total acid content,total phenolic content and color density. Control samples withoutany treatments were also carried out.

Factor level

�ffiffiffi2p

�1 0 +1 þffiffiffi2p

55.9 60 70 80 84.12.9 5 10 15 17.10.012 0.02 0.04 0.06 0.0682.3 4 8 12 13.7


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2.3. Analytical methods

2.3.1. Extraction yieldThe extraction efficiency of the treatment methods was evalu-

ated by using the extraction yield as an index, which was calcu-lated according to the following equation:

Y ¼ m2 � Cm1 � ð100�wÞ � 100 ð1Þ

where Y was the extraction yield (%) of the treatment method, m1

and w were the mass (g) and the moisture (%) of the initial grapemash, respectively; and m2 and C were the mass (g) and the totalsoluble solid content (%) of the obtained grape juice after centrifu-gation, respectively.

To compare the extraction yields obtained from treatmentmethods, extraction enhancement E (%) was calculated accordingto the following equation:

E ¼ Y2 � Y1

Y1� 100 ð2Þ

where Y1 and Y2 were the extraction yields (%) of two comparedtreatment methods.

2.3.2. Relative viscosityRelative viscosity of juice (grel) was determined by using 15 mL

Ostwald viscometer under temperature of 30 �C [31] and was cal-culated as follow:

grel ¼tto

� �qqo

� �ð3Þ

where t and q were the flow time and the specific mass of juice,respectively; to and qo were the flow time and the specific massof distilled water, respectively.

2.3.3. Reducing sugarsReducing sugar content of grape juice was determined by spec-

trophotometric method using 3,5-dinitrosalicylic acid reagent. Thismethod was proposed by Miller [32].

2.3.4. Total acidsTitratable acidity determination, expressed in equivalent of tar-

taric acid content (g/L), was carried out by diluting a 10 mL aliquotof each sample with 90 mL of distilled water and subsequentlytitrating the sample with 0.1 N NaOH to a pH endpoint of 8.1 [33].

2.3.5. Total phenolicsTotal phenolic content of grape juice was determined as by

spectrophotometric method using Folin–Ciocalteu reagent. Thismethod was proposed by Slinkard and Singleton [34].

2.3.6. ColorThe color of grape juice was measured with a Konica Minolta

Colorimeter (CR-410, Osaka Japan). Grape juice was placed onthe light port using a 5 cm diameter plastic dish with cover. Colorparameters were recorded as L* (lightness), a* (redness) and b*(yellowness). The hue angle (h) (h* = arctan b*/a*) and chroma(C) (C = [(a*)2 + (b*)2]0.5) were also calculated [35].

2.4. Statistical analysis

Response surface methodology was used to find out optimalconditions of ultrasound assisted treatment and of combined ultra-sound and enzyme treatment. The experiments were carried outaccording to a central composite design with 2 factors and 5 levels.Table 1 shows independent variables selected for these two treat-


ments. For each factor, an experimental range was based on our re-sults of a preliminary study (unpublished data). Extraction yieldwas the dependent variable. The complete design consisted of 13experimental points including 4 factorial points, 4 axial pointsand 5 center points and the experiment was carried out in a ran-dom order. The software Modde version 5.0 was used to generatethe experimental planning and to process data.

All experiments were performed in triplicate. The experimentalresults obtained were expressed as means ± SD. Mean values wereconsidered significantly different when P < 0.05. Analysis of vari-ance (ANOVA) was performed using the software Statgraphics plus,version 3.2.

3. Results and discussion

3.1. Enzymatic treatment

The enzymatic treatment of grape mash increased the extrac-tion yield as results of Fig. 1. The graphs show that the enzymeconcentration of 0.04%v/v and the treatment time of 40 min werethe appropriate conditions for the enzymatic treatment, which in-creased extraction yield of treated samples approximately 9.2% incomparison with that of the untreated samples. Treatments withhigher enzyme concentration and longer time did not make signif-icant differences in extraction yield.

Pectinase enzymes are known to work on pectic substanceswhich occur as structural polysaccharides in the middle lamellaand primary cell wall. The presence of macerating side-activitiesin the Pectinex Ultra SP-L preparation, such as cellulases and hemi-cellulases would result in a more complete breakdown of the poly-saccharide structure, causing solubilization of the middle lamellaand improving juice extraction.

Our results agreed with conclusions of many previous studieswhich suggested that pectolytic and cellulolytic enzymes could im-prove juice yield of fruit processing such as studies on apple [36],pineapple [37], carrot [29], elderberry [38], and orange [39].

3.2. Sonication treatment

Based on our preliminary investigations (unpublished data), atemperature of 70 �C and a time of 10 min were chosen as the cen-tral conditions of the central composite rotary design (CCRD). Table2 shows extraction yield of each run according to the experimentalplanning.

Multiple regression analysis was performed on the experimen-tal data and the coefficients of the model were evaluated for signif-icance with a Student t-test. All the linear coefficients weresignificant (P < 0.05). One crossproduct coefficient was eliminatedin the refined equation as its effect was not significant. Neglectingthe insignificant term, the final predictive equation obtained is asgiven below:

Y1 ¼ 81:44þ 1:70X1 þ 1:75X2 � 1:60X21 � 1:12X2

2 ð4Þ

where Y1, X1, X2 were the extraction yield of grape mash treatmentby ultrasound (%), the sonication temperature (�C) and the sonica-tion time (min), respectively.

Table 3 presents ANOVA of the fitted model. According to theANOVA table, the regression model is significant at the consideredconfidence level since a satisfactory correlation coefficient was ob-tained and the F-value was 7 times more than the F listed value.

Surface response graph, obtained by using the fitted model pre-sented in Eq. (4), is presented in Fig. 2.

Table 4 presents the estimated effect of each variable, as well astheir interactions on the yield of treatment process. The resultsshow that temperature and time had significantly positive effects


73

74

75

76

77

78

79

80

81

0 0.02 0.04 0.06 0.08 0.1 0.12

Enzyme concentration (%v/v)

Yiel

d (%

)

72

73

74

75

76

77

78

79

80

0 10 20 30 40 50 60 70

Treatment time (min)

Yiel

d (%

)A

B

Fig. 1. Effects of enzyme concentration (A) and treatment time (B) on extractionyield of enzymatic treatment of grape mash.

Table 3Analysis of variance of the regression model in experiments of sonication treatment.

Source of variation SS DF MS F

Regression 76.93 5 14.936 46.423Residual 2.25 7 0.322Total 79.09 12 6.411Listed F-valuea F(4, 4) = 6.4

SS: sum of squares; DF: degrees of freedom; MS: mean square; F: F-value.a F-value at 95% of confidence level.

81.5 .. 82.380.5 .. 81.579.5 .. 80.5

78.5 .. 79.577.5 .. 78.574.0 .. 77.5

Fig. 2. Fitted surface for yield of ultrasound assisted treatment of grape mash as afunction of temperature and time.

Table 4Estimated effect of independent variables on extraction yield of sonication treatment.

Factora Effect Standard error P

X1 3.40162 0.200553 6.26877E-005X2 3.50258 0.200553 5.18922E-005X1 � X1 �3.19049 0.2151 0.000147368X2 � X2 �2.24022 0.2151 0.00124288

X1: sonication temperature, X2: sonication time (min).P indicates significance of linear regressions.

a Significant factors at 95% of confidence level.


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on yield of the treatment process, while their obvious quadratic ef-fects were also observed, but were negative; and temperature hadstronger effect on extraction yield than time.

The enhancement of extraction yield by ultrasound is attributedto a physical phenomenon called acoustic cavitation which in-cludes the formation, growth, and violent collapse of small bubblesor voids in liquids as a result of pressure fluctuation [40]. Collapse


of the bubbles causes shock wave that passes through the solvent,enhancing the mass transfer within the system [5,6]. At high tem-perature, the intensity of bubble collapse is weak by the higher va-por pressure. However, increased temperature augments thenumber of cavitation bubbles as well as decreases the viscosityresulting to a more violent collapse. Thus, there is an optimal tem-perature at which the viscosity is low enough to form enough vio-lent cavitation bubbles, yet the temperature is low enough to avoidthe dampening effect on collapse by a high vapor pressure [41]. Inour study, the optimal temperature of the sample during ultrasonictreatment was about 74 �C (Fig. 2). Our results agreed with previ-ous researches of other authors who reported that sonication at70 �C had positive effect on extraction of some compounds of otherplant materials such as phenolic compounds [9], anthocyanins [7],tartaric and malic acids [10]. The higher temperatures resulted inthe lower extraction yield. Ultrasound has been reported to in-crease the extractability of polysaccharides from plant materials[13,19]. These substances block drainage channels in the pulpthrough which the juice must pass [1]. As a result, the extractionyield was lower.

The optimal time of sonication treatment obtained from Fig. 2was 13 min. Under optimal conditions, the model predicted a max-



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imum response of 82.3%. This value of extraction yield was 12.9%higher than that of the untreated sample. As a result, applicationof ultrasound in grape mash treatment increased the extractionyield 3.4% more than traditionally enzymatic treatment and theprocess time was shortened over three times.

3.3. Combined ultrasound and enzyme treatment

In this experiment, an enzyme concentration of 0.04%v/v and atime of 8 min were chosen as the central conditions of the CCRDaccording to our preliminary results (unpublished data). Table 5shows results of extraction yield for each run obtained from theexperiments.

In order to establish fitted model, multiple regression analysiswas also performed on the experimental data and the final predic-tive equation obtained is as given below:

Y2 ¼ 80:42þ 1:86X3 þ 0:61X4 � 1:36X23 � 0:59X2

4 ð5Þ

where Y2, X3 and X4 were the extraction yield of grape mash treat-ment by combined ultrasound and enzyme method (%), the enzymeconcentration (%v/v) and the treatment time (min), respectively.

The regression model was significant (P < 0.05) because the F-value was 8 times more than the F listed value according to anal-ysis of variance which is presented in Table 6.

In order to determine optimal levels of the variables for theextraction yield of the treatment, three-dimensional surface plotswere constructed according to Eq. (5) (Fig. 3).

According to the estimated effect of each variable as well astheir interactions on the extraction yield in Table 7, change in en-zyme concentration or time resulted in significant change inextraction yield of the treatment process.

From the model, the obtained optimal conditions were the en-zyme concentration of 0.05%v/v and the time of 10 min, at whichthe model predicted a maximum response of 81.2%. This value ofextraction yield was 11.4% higher than that of untreated sample.

Table 6Analysis of variance of the regression model in experiments of combined ultrasoundand enzyme treatment.

Source of variation SS DF MS F

Regression 44.864 5 8.973Residual 1.144 7 0.163 54.927Total 46.008 12 3.834Listed F-valuea F(4, 4) = 6.4

SS: sum of squares; DF: degrees of freedom; MS: mean square; F: F-value.a F-value at 95% of confidence level.

Table 5Experimental planning and results of extraction yield for combined ultrasound andenzyme treatment.

Run Enzyme concentration (%v/v) Time (min) Yield (%)

1 0.02 4 76.02 0.06 4 80.13 0.02 12 77.44 0.06 12 81.25 0.012 8 76.06 0.068 8 81.17 0.04 2.3 79.28 0.04 13.7 81.29 0.04 8 81.110 0.04 8 81.111 0.04 8 80.812 0.04 8 80.913 0.04 8 80.7


As a result, combination of ultrasound and enzyme in grapemash treatment increased extraction yield 2.0% more than tradi-tionally enzymatic treatment and the process time was shortenedover four times; however, its yield was slightly lower than that inthe sonication treatment (Section 3.2). The results of Section 3.2showed that the optimal temperature of the sonication treatmentwas 74 �C while the temperature of 50 �C was kept in this experi-ment to maintain enzyme activity. Consequently, effect of ultra-sound on extraction yield decreased and the extraction yield inthis case was lower. However, the treatment time of this methodwas lower than that of the sonication method.

The understanding of the actual effect of ultrasound on en-zymes is very little because contradictory results of inactivationand activation of enzymes upon ultrasound treatment have beenreported. Unlike traditional heat denaturation, the sonication pro-cess does not destroy all of enzymes [42]. According to Yachmenevet al. [25], when ultrasound was specifically used to inactivate en-zymes, its actual efficiency was quite low and contrary to commonbelief, low intensity and uniform sonication does not damage or

80.5 .. 81.279.5 .. 80.578.5 .. 79.5

77.5 .. 78.576.5 .. 77.575.0 .. 76.5

Fig. 3. Fitted surface for yield of combined ultrasound and enzyme treatment ofgrape mash as a function of enzyme concentration and treatment time.

Table 7Estimated effect of independent variables on yield of ultrasound assisted enzymatictreatment.

Factora Effect Standard error P

X3 3.72842 0.142909 3.62442E-006X4 1.2115 0.142909 0.00384638X3 � X3 �2.72067 0.153274 4.67098E-005X4 � X4 �1.17021 0.153274 0.00656462

X3: enzyme concentration (%v/v), X4: treatment time (min).P indicates significance of linear regressions.

a Significant factors at 95% of confidence level.


79

82

85

88

eld

(%)

Treated sample

Control sample

A


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inactivate sensitive structures of enzyme protein macromolecules[25]. In this study, ultrasound with intensity of 2 W/cm2 improvedthe transport of enzyme macromolecules but does not generate anexcessive amount of high reactive intermediates which cause deac-tivation of enzymes [25]. Moreover, ultrasound was also applied toactivate the catalytic performance of the enzyme macromoleculesadsorbed onto the surface of substrate and to enhance removal ofthe products of hydrolytic reaction from the reaction zone [25].Therefore, ultrasound increased the efficiency of enzymatic treat-ment with higher extraction yield and lower treatment time.

70

73

76

0 0.02 0.04 0.06 0.08 0.1 0.12

Enzyme concentration (%v/v)

Yi88B

3.4. Enzymatic treatment after sonication

As results of Section 3.2, sonication increased extraction yield ofgrape mash treatment, but it also increased content of polysaccha-rides in the treated samples and this phenomenon made difficul-ties for free-run juice recovery. If these substances were brokendown, the extraction yield would be higher. Therefore, we exam-ined enzymatic treatment after sonication using the optimalparameters, i.e. the temperature of 74 �C and the time of 13 min.The results are presented in Fig. 4.

The graphs show that the enzyme concentration of 0.06%v/vand the time of 20 min were the appropriate conditions for theenzymatic treatment after sonication. This treatment increasedthe extraction yield approximately 3.8% more than sonicationtreatment and 7.3% more than enzymatic treatment.

70

73

76

79

82

85

0 10 20 30 40 50

Treatment time (min)

Yiel

d (%

) Treated sample

Control sample

Fig. 4. Effects of enzyme concentration and treatment time on enzymatic treatmentafter sonication.

3.5. Comparison in physico-chemical characteristics of grape juiceobtained from different grape mash treatments

The above results indicated that treatment by ultrasound orcombination of ultrasound and enzyme improved extraction yieldas well as shortened treatment time in comparison with tradition-ally enzymatic treatment of grape mash, and enzymatic treatmentafter sonication made the extraction yield increase more. In thisexperiment, we determined some physico-chemical characteristicsof grape juice obtained from these treatments. The results are pre-sented in Table 8.

Pectinases are able to break down pectin molecules, mainly col-loidal compounds of grape juice. As a result, enzymatic treatment(ET) decreased viscosity of grape juice (Table 8). On the contrary,sonication treatment (ST) with ultrasound wave of 2 W/cm2 inten-sity was not only unable to break down pectin molecules but alsoextracted macromolecules from cell walls which increased viscos-ity of the obtained grape juice. Enzymatic treatment after sonica-tion (ETAS) lowered viscosity due to its ability of pectinbreakdown. However, some other colloidal macromolecules ex-tracted by ultrasound were not broken down by Pectinex UltraSP-L preparation. This was the reason why the viscosity of grapejuice in this method was still higher than that in the enzymatictreatment. In combined ultrasound and enzyme treatment (CUET),enzyme decreased viscosity while ultrasound increased it. Conse-quently, viscosity of grape juice in this method was similar to thatof the control sample.

Table 8 also shows that the content of reducing sugars in ET, ST,CUET and ETAS increased 6.2%, 12.0%, 10.9% and 15.4%, respectivelyin comparison with that in the control sample. Although the differ-ence in extraction yield of ET and ST was low (3.4%), the differencein sugar contents between them was higher (5.8%). The reasoncould be that although ET generated grape juice with lower sugarcontent, it increased volume of the obtained grape juice. Conse-quently, the difference in extraction yield was lower.

With regards to total acid content, Table 8 shows that its valuesin ET, ST, CUET and ETAS increased 9.9%, 13.6%, 10.9% and 14.3%,respectively in comparison with that in the control sample. These


results suggested that ST possessed greater ability of acid extrac-tion than ET.

In comparison with the control sample, all treated samples con-tained significantly higher total phenolic content which increased93.0%, 114.3%, 89.3% and 120.8% in ET, ST, UAET and ETAS, respec-tively. Our results agreed with many previous researches which re-ported that ultrasound possessed high extractability for phenoliccompounds such as anthocyanins [7] and total phenolics [9]. Theresults showed that ST extracted phenolics more effectively thanET. In grape cells, phenolic compounds can link with various com-pounds of cell walls such as polysaccharides or proteins. As a re-sult, random breakdown of cell wall by ultrasound was moreeffective than selective breakdown by enzymes. That was the rea-son why the content of phenolics liberated in the ultrasound treat-ment was higher.

Table 8 also shows that application of all treatment methodsimproved color of the obtained grape juice due to higher values


Table 8Comparison in physico-chemical characteristics of grape juice obtained from different grape mash treatments.

Treatment method Relative viscosity Reducing sugars (g/L) Total acidity (g tartaric acid/L) Phenolics (g/L) C* L* H*

C 1.35 ± 0.01a 122.8 ± 0.5a 4.13 ± 0.01a 2.56 ± 0.01a 50.6 ± 0.5a 24.7 ± 0.7a 57.7 ± 1.0a

ET 1.31 ± 0.01b 130.4 ± 0.3b 4.54 ± 0.01b 4.93 ± 0.03b 49.4 ± 0.9ab 30.7 ± 0.3b 49.0 ± 1.6b

ST 1.67 ± 0.01c 137.5 ± 0.6c 4.69 ± 0.01c 5.48 ± 0.01c 48.4 ± 0.6bc 29.9 ± 0.8bc 43.5 ± 0.7c

CUET 1.38 ± 0.01d 141.8 ± 0.3d 4.72 ± 0.01d 5.64 ± 0.04d 49.0 ± 1.0bc 29.7 ± 0.3c 43.1 ± 0.4c

ETAS 1.35 ± 0.01a 136.1 ± 0.8e 4.58 ± 0.01e 4.84 ± 0.03e 48.1 ± 0.1c 30.0 ± 0.0bc 43.7 ± 0.6c

C: control sample, ET: enzymatic treatment, ST: sonication treatment, CUET: combined ultrasound and enzyme treatment, ETAS: enzymatic treatment after sonication, C*:chroma; L*: lightness; H*: hue angle.Each value is expressed as mean and standard deviation.Values are significantly different (P = 0.05) from other values within a column unless they have at least one similar superscript letter.


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of C* and lower values of H*. It should be noted that ST, CUET andETAS produced grape juices with lower values of H* than ET. Theseresults illustrated that red pigment content of grape juice obtainedfrom ST, CUET and ETAS was higher than that from ET. In otherwords, application of ultrasound in grape mash treatment im-proved color of the obtained grape juice more effectively thanapplication of commercial enzyme. Table 8 also reports that light-ness of all treated samples decreased because of the increase incolor density.

4. Conclusions

In comparison with traditionally enzymatic treatment, applica-tion of ultrasound in grape mash treatment enhanced extractionyield and shortened treatment time. Besides, these methods im-proved quality of the obtained grape juice because they increasedsugar content, total acid content, phenolics content as well as colordensity of grape juice.

References

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Application of Ultrasound in Grape Mash Treatment in Juice Processing