Influence of ultrasonic treatment on the structure and emulsifying properties of PPI

food and bioproducts processing 92(2014)30–37

Contents lists available at ScienceDirect

Food and Bioproducts

Processing

j o u r n a l h o m e p a g e :w w w.e l s e v i e r.c o m /l o c a t e /f b

In?uence of ultrasonic treatment on the structure and emulsifying properties of peanut protein isolate

Qiu-Ting Zhang a ,Zong-Cai Tu a ,b ,?,Hui Xiao c ,Hui Wang d ,e ,??,Xiao-Qin Huang b ,Guang-Xian Liu d ,Cheng-Mei Liu a ,Yan Shi a ,Liang-Liang Fan a ,De-Rong Lin f

State Key Laboratory of Food Science and Technology,Nanchang University,Nanchang,Jiangxi,330047,China b

Jiangxi Normal University,Nanchang,Jiangxi,330022,China

c Albert Einstein College of Medicine,Yeshiva University,Bronx,NY 10461,Unite

d States

d Engineering Research Center for Biomass Conversion,Ministry of Education,Nanchang University,Nanchang,Jiangxi,330047,China

e Food Research Centre,Jiangxi Academy o

f Agricultural Sciences,Nanchang,Jiangxi,330200,China

f Harbin Inst Technol,State Key Lab Urban Water Resource &Environment,Harbin,Heilongjiang,150090,China

a b s t r a c t

Effects of ultrasonic treatment on emulsifying properties and structure of peanut protein isolate (PPI)were evalu-ated by analysis of particle size distribution,protein surface hydrophobicity,SDS-PAGE,circular dichroism spectra

and environmental scanning electron microscopy.The emulsifying properties of the PPI were found to be improved

by ultrasonic treatment.The mean particle size decreased from 474.7nm to 255.8nm while the molecular weight

remained unaffected.The results of intrinsic ?uorescence spectroscopy and surface hydrophobicity indicated that

ultrasonic treatment induced tertiary structural changes of the proteins in PPI.Emulsifying activity index and emul-sion stability index were found to be correlated fairly well with surface hydrophobicity (H 0)(r =0.712and r =0.668,

respectively).

Crown Copyright ?2013Published by Elsevier B.V .on behalf of The Institution of Chemical Engineers.All rights

reserved.

Keywords:Ultrasonic treatment;Peanut protein isolate;Emulsifying properties;Structure;Surface hydrophobicity

Introduction

Peanut protein isolate (PPI)is the by-product of the peanut oil squeezing process in China.The output of defatted peanut ?our is large,and it contains 47–55%proteins with high nutri-tion value (Basha and Pancholy,1982).However,these proteins still remain underutilized (Yu et al.,2007).The functional prop-erties of PPI,i.e.,emulsifying,foaming and gel properties are not as good as soy protein isolate (Wu et al.,2009).Although PPI has good potential to be a source of nutritional proteins,its poor functional properties have greatly limited its application in the food industry.

Numerous methods have been investigated to improve protein’s functional properties,including heat treatments,enzymatic hydrolysis and high-pressure homogenization

Corresponding author at :State Key Laboratory of Food Science and Technology,Nanchang University,235Nanjing East Road,Nanchang 330047,Jiangxi,China.Tel.:+008679188305938;fax:+008679188305938.??

Corresponding author at :Engineering Research Center for Biomass Conversion,Ministry of Education,Nanchang University,235Nanjing East Road,Nanchang 330047,Jiangxi,China.Tel.+008679188305938.

E-mail addresses:T uzc mail@https://www.360docs.net/doc/499171280.html, (Z.-C.T u),wanghui00072@https://www.360docs.net/doc/499171280.html, (H.Wang).Received 18October 2012;Received in revised form 15July 2013;Accepted 19July 2013

(Chen et al.,2011;Penas et al.,2004;Sorgentini et al.,1995).Ultrasonic treatment is frequently used in chemical synthe-sis,preparation of pharmaceuticals,polymer,chemical,textile and cosmetic industries (Canselier et al.,2002;García-Pérez et al.,2007;Noshad et al.,2012).Recently,it has been used in food processing for changing viscosity and texture,func-tionality of dairy products,meat tenderization,mixing and emulsi?cation (Jayasooriya et al.,2004),degassing and foam control (Villamiel et al.,2000)and improving the extraction process of food bioactive substances (Vilkhu et al.,2008).It has also been applied in other ?elds,for example,food drying,assisted crystallization and sterilization (De la Fuente-Blanco et al.,2006;Luque de Castro and Priego-Capote,2007;Villamiel et al.,2000).Ultrasonic treatment not only represents a rapid,ef?cient and reliable alternative to improve the quality of food,

https://www.360docs.net/doc/499171280.html,/10.1016/j.fbp.2013.07.006

food and bioproducts processing92(2014)30–3731

but also has the potential to develop new products with a unique functionality(Soria and Villamiel,2010).

In the present work,we?rst prepared PPI using alkali extraction followed by acid precipitation.We then evaluated the effect of ultrasonic treatment on the emulsifying proper-ties of PPI.The emulsifying properties of PPI were found to be correlated with the protein structure.

2.Materials and methods

2.1.Materials

Peanut protein powder(80%protein,7.3%moisture,3.2%fat and6%ash)(Baiaote plant protein science technology Co.,Ltd., Shanghai,China),bovine serum albumin(BSA),5,5 -dithiobis-(2-nitrobenzoic acid)(DTNB),8-anilinonaphthalene-sulfonic acid(ANS),were purchased from Sigma Chemical Co.(St, Louis,MO,USA).Sodium dodecyl sulphate(SDS),Coomassie Blue R-250and N,N,N ,N -tetramethyl ethylene diamine (TEMED),blue prestained low molecular weight protein marker(14.4–97.4kDa)were obtained from Beijing Solarbio Science&Technology Co.,Ltd.(Beijing,China).Soybean oil was purchased from a local supermarket and used directly without further puri?cation.All chemicals used in the present study were of analytical grade.

2.2.Preparation of peanut protein isolate

Peanut protein powder was suspended in12-fold distilled water and the pH was adjusted to8.0with2N NaOH at50?C for1h.After centrifugation at4000r/min for20min,the pH of the supernatant was adjusted to4.5with2N HCl to precipitate the peanut proteins.The precipitated proteins were collected and dissolved in distilled water.The pH was adjusted to7.0 followed by dialysis with distilled water at4?C for24h.After freeze-drying,the peanut proteins were stored in4?C until their use in ultrasound modi?cation and other tests.

2.3.Ultrasound pretreatment of PPI

PPI dispersions(8g L?1)were prepared by adding PPI pow-der into distilled water and then gently stirred overnight at ambient temperature(25?C).The PPI dispersions were treated by ultrasonic cell disruption instrument(JY98-IIIDN,Ningbo Xinzhi Instruments,Inc.,Ningbo,China,20kHz)with a2.0cm diameter titanium probe.100mL of PPI dispersions in150mL ?at bottom conical?asks were immersed in a low temper-ature thermostat(DC-2010,Ningbo Xinzhi Instruments,Inc., Ningbo,China),which can control the temperature ranging from?20?C to100?C.Samples were treated under different levels of power output(0,120,300,480,660,840,1020W)at different temperatures(0,15,25,35,45,55,65,75?C)for0,1, 3,5,10,20,30min(pulse durations of on-time9s and off-time 1s),respectively.After freeze-drying,the samples were stored in4?C until further tests.

2.4.Determination of emulsifying properties

Emulsifying properties were determined according to the method of Jamdar et al.(2010)with a little modi?cation. Soybean oil(10mL)and30mL of1%protein solution were mixed.The mixture was homogenized using a homogenizer (Ika-Ultra-T ur-rax T25,Germany)at a speed of10,000rpm for 2min.An aliquot of the emulsion(50?L)was pipetted from the bottom of the container at0and10min after homogenization and mixed with5mL of0.1%SDS solution.The absorbance of the diluted solution was measured at500nm using a spec-trophotometer(T6,Pgeneral,Beijing,China).The absorbance measured immediately(A0)and10min(A10)after emulsion formation were used to calculate the emulsifying activity index(EAI)and emulsion stability index(ESI)as follows:

EAI(m2/g)=

(2×2.303×A0)

[0.25×protein weight(g)]

ESI(min)=A0× t/ A

where A10is the absorbance at10min after homogenization; t=10min;and A=A0–A10.

2.5.Determination of mean particle size and distribution(PSD)

The mean particle size and particle size distributions(PSDs) were determined with a by Nicomp380/ZLS Zeta poten-tial/Particle sizer(PSS Nicomp,Santa Barbara,USA)according to the methods of Liu et al.(2011a).The Nicomp380,based on dynamic light scattering(DLS),provides accurate mean parti-cle size and PSD down to1nanometer.Samples were diluted by approximately1/1000with deionized water in the sample dispersion unit under stirring.

2.6.SDS-PAGE

SDS-PAGE was performed according to the method of Laemmli (1970)with slight modi?cations.A12%acrylamide separat-ing gel and a5%acrylamide stacking gel containing0.1%SDS were used in a Bio-rad Mini protean Tetra MP4electrophoresis (Bio-Rad Mini-Protean System).The samples were mixed with loading buffer(containing0.06M Tris–HCl buffer(pH8.8),con-taining2%SDS,5%?-mercaptoethanol,25%glycerol and0.1% bromophenol blue).The solutions were then heated in boiling water for10min and centrifuged at10,000g for3min before electrophoresis.Ten microliters of each sample were loaded onto the gel.Low molecular weight markers(14.4–97.4kDa) were run as a reference.Electrophoresis was performed in electrophoresis buffer(containing0.025M Tris–HCl,0.192M Glycine and0.1%SDS)for30min at16mA,followed by1.5h at32mA.Gels were stained with Coomassie Blue G250and destained in a20%ethanol/80%deionized water mixture. 2.7.Fluorescence spectra analysis

2.7.1.Intrinsic?uorescence

The intrinsic emission?uorescence spectra of the protein samples were obtained by a Fluor photometer(F4500,Hitachi, Tokyo,Japan)according to Liu et al.(2011b)with modi?cations. Protein solutions(0.2mg mL?1)were prepared in10mM phos-phate buffer(pH7.0).To minimize the contribution of tyrosine residues to the emission spectra,the protein solutions were excited at290nm,and emission spectra were recorded from 300to460nm at a constant slit of2.5nm for both excitation and emission.All the measurements were conducted in trip-licate.

2.7.2.Surface hydrophobicity(H0)Measurement

H0values of samples were determined using ANS as the?u-orescence probe according to the method of Kato and Nakai (1980).Lyophilized PPI and UPPI samples(1.0mg mL?1in0.01M phosphate buffer at pH7.0),and serially diluted with the same

32food and bioproducts processing92(2014)30–37

buffer to obtain protein concentrations ranging from1.0to 0.02mg mL?1.Then20?L of ANS(8.0mM in0.01M phosphate buffer,pH7.0)was added to2mL of the sample.The relative ?uorescence intensities of the ANS-proteins(native PPI and UPPI)conjugates were measured at room temperature using a?uorescence spectrophotometer.Fluorescence intensity(FI) was measured with a Hitachi F4500?uorescence spectrometer (LS55)at wavelengths of390nm(excitation)and470nm(emis-sion),with a constant excitation and emission slit of5nm.The protein hydrophobicity was expressed as the initial slope of relative?uorescence intensity(RFI)vs.protein concentration (mg mL?1)(calculated by linear regression analysis).

2.8.Circular dichroism(CD)spectra analysis

Protein secondary structure was determined using CD accord-ing to Jacks et al.(1973).Slight modi?cation of Green?eld and Fasman(1969)was applied.A MOS-450circular dichroism spectrometer equipped with a Peltier element(Bio-Logic Sci-ence Instruments,Grenoble,France)was used for CD analysis. The far-UV CD spectroscopic measurements were done in a 2mm path length quartz cuvette with a sample concentration of0.1mg mL?1.All of the protein samples were centrifuged at 12,000g for20min at20?C,prior to the analysis of the super-natant.The samples were scanned from190to250nm,with a scan rate of50nm min?1.A mean value of148g mol?1for amino acid residue was assumed in all calculations,and CD measurements were expressed as mean residue ellipticity(?, degree cm2dmol?1).The secondary structure compositions of the samples were calculated from the mean residue ellipticity using CD Pro software(Bio-Logic Science Instruments)with the CONTIN/LL program assuming small peptide structure parameters are the same as those for proteins.

2.9.Scanning microscopy analysis

PPI and UPPI samples were taken after freeze-drying,and sam-ples were prepared by sticking the protein onto double-sided adhesive tape attached to a circular specimen stub.The sam-ples were viewed using an environmental scanning electron microscope(ESEM)(Quanta200F,FEI,Deutschland GmbH,Kas-sel,Germany)at30kV and3.0spot size.The low vacuum mode was used while operating the ESEM.

2.10.Statistical analysis

All the experiments were done in triplicate.The data were expressed as mean±standard deviation(SD).One-way analy-sis of variance(ANOVA)and Pearson’s Bivariate Correlation test were carried out to calculate statistical signi?cance in addition to their correlation coef?cients(r)using analytical software SPSS statistics(Version17.0,SPSS Inc.,Chicago,IL, USA).

3.Result and discussion

https://www.360docs.net/doc/499171280.html,position of PPI

The proximate composition of PPI is displayed in Table1.The protein content was determined to be91.60%,similar to the reports by Wu et al.(2009)and Yu et al.(2007).The content of crude fat was decreased to0.76%by the alkali solvation and isoelectric precipitation;the ash content of PPI was mea-sured to be2.88%.Sosulski and McCurdy(1987)indicated that

Table1–Proximate analyses.

Proximate compositions a(%wet wt)

Moisture Protein Crude fat Ash

PPI 4.4±0.191.6±0.40.76±0.04 2.9±0.1

a Each value is expressed as mean±standard deviation.

strong acid and alkali used in pH adjustment during protein extraction may contribute to salt formation and results in high values for ash content in the products.

3.2.Emulsifying properties

Emulsifying properties express the interfacial area stabilized per unit weight of a protein,characterizing the ability of a protein to absorb to the oil-water interface.Emulsifying prop-erties of food proteins can be generally evaluated by the EAI and ESI(Jamdar et al.,2010).Fig.1(A)showed the effects of ultrasound time on the EAI and ESI values of PPI solutions under600W at20?C.The lowest EAI value(18.6m2g?1)was obtained at the native PPI while the EAI value was increased signi?cantly after just1min ultrasonic treatment.No signif-icant difference was observed with longer treatment.ESI of PPI showed the same trend as EAI.Fig.1(B)showed the EAI and ESI values of PPI samples as function of ultrasound power at5?C for5min.The EAI values were increased signi?cantly when the ultrasound power was higher than660W.When the ultrasound power was higher than120W,the ESI value of UPPI was much higher than that of the native PPI.No signi?-cant difference showed after the power higher than120W.The results were consistent with the report in which high power ultrasound modi?ed the functional properties of protein such as solubility and emulsifying properties(Karki et al.,2009). Fig.1(C)showed the EAI and ESI of PPI treated by different ultrasound temperatures with660W for5min.The values of EAI and ESI were increased with the ultrasound temperature increasing.The highest EAI(32.2m2g?1)of UPPI was obtained at75?C,which was much higher than that of the native PPI. ESI of PPI showed the same trend as EAI under various tem-peratures.

The emulsifying properties of protein were related to the protein solubility,surface charge,surface hydrophobicity and molecular?exibility.Kreˇs i′c et al.(2008)reported that ultra-sound can modify the functional properties of whey protein with some secondary structure changes.The effects of ultra-sound are a combination of thermal,mechanical and chemical effects(Crum,1995;Hu et al.,2012;Mason and Cordemans, 1996;Yu et al.,2007).High temperature may affect the emul-sifying properties by changing the particle size,secondary structure,and surface hydrophobicity of proteins.Proteins will unfold with interrupted hydrophobic interactions and disul-?de formation(Yu et al.,2007).The denaturation temperature of PPI is higher than90?C,so the ultrasound temperature(up to75?C)should not be the factor that induces the denaturation of proteins.The instantaneous extreme temperatures and pressures generated by ultrasound might be the major force that changing the emulsifying properties of the PPI.It should be noted that the denatured proteins will expose hydrophobic groups,which usually leading to protein aggregation.There-fore,there is a balance between the aggregation and exposure of hydrophobic groups(Soria and Villamiel,2010).

food and bioproducts processing92(2014)30–37

Fig.1–Emulsifying properties of PPI in different ultrasound time at600W,20?C(A),powers at5?C,5min(B) and temperatures at660W,5min(C).Error bars show mean standard deviation of three determinations.

3.3.Particle size

The mean particle size and particle size distribution(PSD)of PPI solutions(PBS)under different ultrasonic treatments were shown in https://www.360docs.net/doc/499171280.html,pared to the native PPI,the mean parti-cle size of PPI with ultrasound power above120W,decreased signi?cantly(Fig.2(A)).Ultrasonic treatment was not affected by higher temperature as shown in Fig.2(B).Longer treat-ment time resulted in a further decreased particle size of PPI (Fig.2(C)).The PSD of UPPI and PPI was determined as shown in Fig.2(D).The mean particle size of native PPI is474.7nm and the range of its particle size is from50nm to2.5?m.After ultrasonic treatment,the mean particle size was decreased to255.8nm,and the range of its particle size is from20nm to1?m.It was shown that PSD of samples was signi?cantly affected by ultrasonic treatment.As shown in Fig.2(D),the distribution of particles was much more centralized by the ultrasound treatment with much narrower distribution width.

3.4.SDS-PAGE

SDS-PAGE pro?les of the samples are shown in Fig.3.The elec-trophoresis pro?le of peanut protein subunits contains?ve main classes(de Jong et al.,1998).Comparing to the native PPI,ultrasonic treatment did not induce major changes in the protein electrophoresis pro?les,indicating that ultrasonic treatment did not change the primary structure of the pro-teins.This is consistent with the previous observation that no dimerization was formed after ultrasonic treatment condi-tions(Chen et al.,2011;Zhao et al.,2011).Similar results have also been obtained in the SDS-PAGE studies of soybean protein isolate(Hu et al.,2012;Karki et al.,2009).

3.5.Intrinsic?uorescence emission spectroscopy analysis

The intrinsic?uorescence is dependent on the polarity of the environment of the tryptophan(Trp)/tyrosine(T yr)residues or Trp/T yr speci?c interactions.When a protein unfolds,the chro-mophores become exposed to solvent,resulting in a decreased ?uorescence intensity(Pallarès et al.,2004).At280nm,both Trp and T yr residues of a protein will be exicited,the changes of tertiary structure of the protein can therefore be deter-mined from the?uctuations of the?uorescence(Liu et al., 2010;Shen and Tang,2012).Intrinsic?uorescence emission spectra of native PPI and UPPI are presented in Fig.4.With dif-ferent ultrasonic treatments,the wavelength of?uorescence emission peak was not shifted.However,the protein clearly exhibited decreased relative?uorescence intensity under all ultrasonic treatments.A reasonable explanation for this is that the protein structure was disrupted by ultrasonic treat-ment,causing more chromophores exposed to the solvent; therefore,the?uorescence intensity was decreased.

3.6.Surface hydrophobicity

Protein surface hydrophobicity(H0)was an index of the num-ber of hydrophobic groups on the surface of a protein in contact with the polar aqueous environment and closely related to its emulsifying properties.The surface hydropho-bicity exhibited by?uorescence emission spectra of PPI and UPPI are shown in Fig.5(A).UPPI showed signi?cantly elevated ?uorescence intensity,suggesting that the surface hydropho-bicity was increased upon protein unfolding.Similar to our results,ultrasonic treatment has been reported to increase the surface hydrophobicity of SPI(Chen et al.,2011;Hu et al., 2012).In addition,as shown in Fig.5(B),the H0value of UPPI increased with elevated temperature.When the temperature was increased to75?C,the H0value was drastically higher than that of other temperatures,suggesting that temperature plays an important role in affecting the surface hydrophobic-ity.It is possible that more hydrophobic regions of PPI buried inside of the protein became exposed to the surface.

H0,EAI and ESI of PPI treated by orthogonal experiment and the correlation analysis were summarized in Tables2and3. The correlation coef?cients are r=0.712and r=0.668for H0

34food and bioproducts processing92(2014)30–37

Fig.2–The mean particle size of PPI in ultrasound power at5?C,5min(A),temperature at660W,5min(B),time at600W, 20?C(C).Error bars show mean standard deviation of three determinations and particle size distribution(PSD)of native PPI and UPPI.

Fig.3–Sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE)pro?les of PPI.(M,molecular weight marker; native PPI,Lane1;15min–660W?25?C,Lane2;15min–720W?30?C,Lane3;20min–600W?25?C,Lane4;20min–660W?30?C,Lane5;20min–720W?20?C,Lane6;25min–600W?30?C,Lane7;25min–660W?20?C,Lane8;25min–720W?25?C,Lane9).

food and bioproducts processing 92(2014)30–37

F l u o r e s c e n c e i n t e n s i t y (A .U )

Wavelength (nm)

Fig.4–Intrinsic ?uorescence intensity of PPI and UPPI (1.Native;2.15min –720W ?30?C;3.20min –600W ?25?C;4.20min –660W ?30?C;5.20min –720W ?20?C;6.25min –600W ?30?C).

vs.EAI,and H 0vs.ESI,respectively,suggesting that there are fairy strong correlations between H 0and EAI/ESI.This is con-sistent with the previous research results (Voutsinas et al.,1983).Since ultrasonic treatment can induce protein unfol-ding,exposing hydrophobic group of proteins,it can be used to improve emulsifying properties of PPI.

3.7.

Circular dichroism spectra

Hu et al.(2012)reported that high power ultrasonic treatment (400W ,30min and 600W ,30min)can induce an increase in ?-helix and random coil,and a decrease in ?-sheet of soybean protein.Low power (200W ,15min and 200W 30min)showed a decrease in ?-helix and random coil,and a increase in ?-sheet of soybean protein.Gülseren et al.(2007)found a small decrease in random coil content of bovine serum albumin (BSA)after ultrasonic treatment (20kHz,450W)with longer ultrasound time.Liu et al.(2011b)had reported ?-helix,?-strand,?-turns and random coil of the native PPI were 7.3%,30.1%,19.4%and 43.4%.This result indicated that the sec-ondary structures of PPI might be affected by ultrasound time,power and temperature.The secondary structure composition of dispersions of PPI and UPPI with different ultrasonic treat-ment by circular dichroism spectra were shown in Fig.6.No

)

U .A (y t i s n e d n i e c n e c s e r o u l F Wavelength (nm)

0 15

1000

1500

2000

2500

3000

H 0

Temperature (oc)

Fig.5–(A)Fluorescence spectra of ANS of PPI and UPPI (1.Native;2.15min –720W ?30?C;3.20min –600W ?25?C;4.20min –660W ?30?C;5.20min –720W ?20?C).(B)Surface hydrophobicity (H 0)of PPI treated by different ultrasonic treatments.

Table 3–Correlations of H 0and EAI/ESI.

EAI

ESI

H 0

Pearson correlation 0.712*0.668*Signi?cant (2sides)0.0210.035N

Correlations signi?cant at the 0.05(2sides).

Table 2–Surface hydrophobicity (H 0),EAI and ESI of PPI treated by ultrasonic treatment orthogonal experiment.

Ultrasound time (min)

Ultrasound power (W)

Ultrasound temper-ature (?C)

H 0EAI

ESI

000

0 1.77×10318.6±0.0212.0±0.0311(15)1(600)1(20) 2.11×10326.9±0.0112.4±0.05212(660)2(25) 2.23×10326.5±0.0111.4±0.03313(720)3(30) 2.51×10329.3±0.0412.0±0.0342(20)12 2.50×10330.3±0.0313.1±0.015223 2.79×10328.1±0.0211.9±0.046231 2.00×10327.3±0.0112.2±0.0173(25)13 3.23×10330.1±0.0314.5±0.048321 2.57×10328.2±0.0113.6±0.049

3.07×103

28.8

0.03

13.3

0.05

food and bioproducts processing 92(2014)30–37

Utrasonic Power (W)

Temperature(oC)

Time (min)

P e r c e n t c o n t e n t (%)

-Helix -Strand -Turns Unordered

Fig.6–Secondary structure distribution of PPI ultrasonic power (A),temperature (B)and time (C),estimated from circular dichroism spectra in the far-UV region at 25?C.

signi?cant difference was found under different conditions,indicating that the secondary structure of PPI was not affected signi?cantly.Therefore,the major contribution for the struc-tural changes of PPI is from tertiary structure but not from secondary structure.

3.8.Environment scanning electron microscopy (ESEM)

Fig.7showed the ESEM images of a magni?cation factor of 3000-fold to evaluate the lyophilized PPI and UPPI by ESEM.Heterogeneous particles consisting of clumps in different sizes and shapes were observed.

After treatment of ultra-sound,the image of the UPPI showed signi?cant difference from that of the native PPI.The surfaces of UPPI were full of holes while the untreated PPI exhibited a more integrated form.It seems that during processing the ultrasonic cavitation effects can force the globular structure into mesh structure.

Conclusions

The emulsifying properties of PPI were found to be improved by ultrasonic treatment.SDS-PAGE,CD and ?uorescence spec-troscopy were used to probe the possible reason that improve emulsifying properties of PPI.We found that it is the ter-tiary structure,but not the primary structure and secondary structure that affect the emulsifying properties.The surface hydrophobicity is found to be well related to the emulsifying properties of PPI.

Fig.7–ESEM micrograph of native-PPI (A)and UPPI (B,25min –600W ?30?C;C,25min –660W ?20?C).

food and bioproducts processing92(2014)30–3737

Acknowledgements

The authors gratefully acknowledge the?nancial support of the National Key Basic Research Program of China (No.2012CB126300),National High Technology Research and Development Program of China(863Program)(No. 2011AA100800),Ganpo Elite555Project of Jiangxi province, and Freedom explore Program of State Key Laboratory of Food Science and Technology of Nanchang University(No.SKLF-TS-200923).

References

Basha,S.M.,Pancholy,S.K.,https://www.360docs.net/doc/499171280.html,position and characteristics of basic proteins from peanut(Arachis hypogaea L.)seed.J.

Agric.Food Chem.30,1176–1179.

Canselier,J.,Delmas,H.,Wilhelm,A.,Abismail,B.,2002.

Ultrasound emulsi?cation–an overview.J.Dispers.Sci.

Technol.23,333–349.

Chen,L.,Chen,J.,Ren,J.,Zhao,M.,2011.Effects of ultrasound pretreatment on the enzymatic hydrolysis of soy protein

isolates and on the emulsifying properties of hydrolysates.J.

Agric.Food Chem.59,2600–2609.

Crum,L.A.,https://www.360docs.net/doc/499171280.html,ments on the evolving?eld of

sonochemistry by a cavitation physicist.Ultrason.Sonochem.

2,S147–S152.

De la Fuente-Blanco,S.,Riera-Franco de Sarabia,E.,

Acosta-Aparicio,V.,Blanco-Blanco,A.,Gallego-Juárez,J.,2006.

Food drying process by power ultrasound.Ultrasonics44,

e523–e527.

Gülseren,˙I.,Güzey,D.,Bruce,B.D.,Weiss,J.,2007.Structural and functional changes in ultrasonicated bovine serum albumin solutions.Ultrason.Sonochem.14,173–183.

García-Pérez,J.V.,Cárcel,J.A.,Benedito,J.,Mulet,A.,2007.Power ultrasound mass transfer enhancement in food drying.Food and Bioprod.Process.85,247–254.

Green?eld,N.J.,Fasman,G.D.,https://www.360docs.net/doc/499171280.html,puted circular dichroism spectra for the evaluation of protein conformation.Biochem.

8,4108–4116.

Hu,H.,Wu,J.,Li-Chan,E.C.Y.,Zhu,L.,Zhang,F.,Xu,X.,Fan,G., Wang,L.,Huang,X.,Pan,S.,2012.Effects of ultrasound on

structural and physical properties of soy protein isolate(SPI) dispersions.Food Hydrocolloid.30,647–655.

Jacks,T.,Barker,R.,Weigang Jr.,O.,1973.Conformations of oilseed storage proteins(globulins)determined by circular

dichroism.Int.J.Pept.Protein Res.5,289–291.

Jamdar,S.,Rajalakshmi,V.,Pednekar,M.,Juan,F.,Yardi,V., Sharma,A.,2010.In?uence of degree of hydrolysis on

functional properties,antioxidant activity and ACE inhibitory activity of peanut protein hydrolysate.Food Chem.121,

178–184.

Jayasooriya,S.,Bhandari,B.,Torley,P.,D‘Arcy,B.,2004.Effect of high power ultrasound waves on properties of meat:a review.

Int.J.Food Prop.7,301–319.

de Jong,E.C.,Van,Z.M.,Spanhaak,S.,Koppelman,S.J.,Pellegrom,

H.,Penninks,A.H.,1998.Identi?cation and partial

characterization of multiple major allergens in peanut

proteins.Clin.Exp.Allergy28,743–751.

Karki,B.,Lamsal,B.P.,Grewell,D.,Pometto,A.L.,Van Leeuwen,J., Khanal,S.K.,Jung,S.,2009.Functional properties of soy

protein isolates produced from ultrasonicated defatted soy

?akes.J.Am.Oil Chem.Soc.86,1021–1028.

Kato,A.,Nakai,S.,1980.Hydrophobicity determined by a ?uorescence probe method and its correlation with surface properties of proteins.Biochim.Biophys.Acta-Protein Struct.

624,13–20.Kreˇs i′c,G.,Lelas,V.,Jambrak,A.R.,Herceg,Z.,Brnˇci′c,S.R.,2008.

In?uence of novel food processing technologies on the

rheological and thermophysical properties of whey proteins.J.

Food Eng.87,64–73.

Laemmli,U.K.,1970.Cleavage of structural proteins during the assembly of the head of bacteriophage T4.Nature227,

680–685.

Liu,C.M.,Zhong,J.Z.,Liu,W.,T u,Z.C.,Wan,J.,Cai,X.F.,Song,X.Y., 2011a.Relationship between functional properties and

aggregation changes of whey protein induced by high

pressure micro?uidization.J.Food Sci.76,E341–E347.

Liu,W.,Zhang,Z.-Q.,Liu,C.-M.,Xie,M.-Y.,T u,Z.-C.,Liu,J.-H., Liang,R.-H.,2010.The effect of dynamic high-pressure

micro?uidization on the activity,stability and conformation of trypsin.Food Chem.123,616–621.

Liu,Y.,Zhao,G.,Zhao,M.,Ren,J.,Yang,B.,2011b.Improvement of functional properties of peanut protein isolate by conjugation with dextran through Maillard reaction.Food Chem.131,

901–906.

Luque de Castro,M.,Priego-Capote,F.,2007.Ultrasound-assisted crystallization(sonocrystallization).Ultrason.Sonochem.14, 717–724.

Mason,T.,Cordemans,E.,1996.Ultrasonic intensi?cation of chemical processing and related operations:a review.Chem.

Eng.Res.Des.74,511–516.

Noshad,M.,Mohebbi,M.,Shahidi,F.,Mortazavi,S.A.,2012.Effect of osmosis and ultrasound pretreatment on the moisture

adsorption isotherms of quince.Food Bioprod.Process.90,

266–274.

Pallarès,I.,Vendrell,J.,Avilés,F.X.,Ventura,S.,2004.Amyloid ?bril formation by a partially structured intermediate state of ?-chymotrypsin.J.Mol.Biol.342,321–331.

Penas,E.,Préstamo,G.,Gomez,R.,2004.High pressure and the enzymatic hydrolysis of soybean whey proteins.Food Chem.

85,641–648.

Shen,L.,Tang,C.-H.,2012.Micro?uidization as a potential technique to modify surface properties of soy protein isolate.

Food Research International48,108–118.

Sorgentini,D.A.,Wagner,J.R.,Anon,M.C.,1995.Effects of thermal treatment of soy protein isolate on the characteristics and

structure-function relationship of soluble and insoluble

fractions.J.Agric.Food Chem.43,2471–2479.

Soria,A.C.,Villamiel,M.,2010.Effect of ultrasound on the technological properties and bioactivity of food:a review.

Trends Food Sci.Tech.21,323–331.

Sosulski,F.,McCurdy,A.,1987.Functionality of?ours,protein fractions and isolates from?eld peas and faba bean.J.Food Sci.52,1010–1014.

Vilkhu,K.,Mawson,R.,Simons,L.,Bates,D.,2008.Applications and opportunities for ultrasound assisted extraction in the

food industry–A review.Innov.Food Sci.Emerg.Technol.9,

161–169.

Villamiel,M.,Verdurmen,R.,Jong,P.,2000.Degassing of milk by high-intensity https://www.360docs.net/doc/499171280.html,chwissenschaft55,123–125. Voutsinas,L.P.,Cheung,E.,Nakai,S.,1983.Relationships of hydrophobicity to emulsifying properties of heat denatured proteins.J.Food Sci.48,26–32.

Wu,H.,Wang,Q.,Ma,T.,Ren,J.,https://www.360docs.net/doc/499171280.html,parative studies on the functional properties of various protein concentrate

preparations of peanut protein.Food Res.Int.42,343–348. Yu,J.,Ahmedna,M.,Goktepe,I.,2007.Peanut protein concentrate:production and functional properties as affected by processing.Food Chem.103,121–129.

Zhao,G.,Liu,Y.,Zhao,M.,Ren,J.,Yang,B.,2011.Enzymatic hydrolysis and their effects on conformational and functional properties of peanut protein isolate.Food Chem.127,

1438–1443.