structuralandphysicochemicalcharacteristicsofgranularmalic … · 2018. 9. 20. · ds 162 ×n naoh...

Research ArticleStructural and Physicochemical Characteristics of GranularMalicAcid-Treated Sweet Potato Starch Containing Heat-StableResistant Starch

Chinwoo Kwon1 Ha Ram Kim1 Tae Wha Moon12 Seung Hyun Lee 3

and Chang Joo Lee 4

1Department of Agricultural Biotechnology Seoul National University Seoul Republic of Korea2Center for Food and Bioconvergence and Research Institute of Agriculture and Life Sciences Seoul National UniversitySeoul 08826 Republic of Korea3Department of Biosystems Machinery Engineering Chungnam National University Daejeon 34134 Republic of Korea4Department of Food Science and Biotechnology Wonkwang University Iksan Jeonbuk 54538 Republic of Korea

Correspondence should be addressed to Chang Joo Lee cjleewkuackr

Received 20 September 2018 Revised 7 December 2018 Accepted 24 December 2018 Published 23 January 2019

Guest Editor Toshinori Shimanouchi

Copyright copy 2019 Chinwoo Kwon et al is is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work isproperly cited

is study investigated the structural and physicochemical characteristics of malic acid-treated sweet potato starch Sweet potatostarch mixed with various concentrations of malic acid solution underwent either thermal or nonthermal treatment Observationof samples under a light microscope ensured the maintenance of granular shape and the Maltese cross FT-IR spectra displayed adistinct carbonyl peak at 1722 cmminus1 and analysis of the degree of substitution (DS) indicated an increase in the extent of esterbonds with increasing concentrations of malic acid e DS of 20M-130 (0214) was the highest and that of 05M-130 was thelowest (0088) among the reacted starches In vitro digestion test revealed an increased amount of resistant starch when a highconcentration of malic acid was used In addition thermally treated samples maintained a higher content of resistant starch (RS)after 30min of cooking at 100degC After cooking 20M-130 had an RS fraction of 534 which was reduced to 499 after cookingrevealing greater heat stability compared with nonthermally treated samples e structure of malic acid-treated starch wasinvestigated using a differential scanning calorimeter (DSC) an X-ray diffractometer a rapid visco analyzer (RVA) and analysis ofapparent amylose content e results showed that thermal and malic acid treatment of starch caused not only partial hydrolysisbut also rearrangement of the crystalline area and helix structure of starch by esterification Analysis of malic acid-treated starchusing a rapid visco analyzer showed no pasting properties due to lack of its swelling caused by the malic acid cross link

1 Introduction

Starch is used in many kinds of food and serves as a majorsource of energy for humans For nutritional purposesstarch can be classified into three categories based on therate of its enzymatic digestion rapidly digestible starch(RDS) slowly digestible starch (SDS) and resistant starch(RS) [1] RS is also defined as the sum of starch and de-graded starch products that resist digestion in the smallintestine of healthy people [2] e RS content of starch isaffected by the amyloseamylopectin ratio physical form

degree of gelatinization storage and thermal treatment[3 4] Depending on the cause of resistance RS can befurther divided into five categories [5 6] RS1 physicallyinaccessible starch due to entrapment RS2 raw starchgranules with crystallinity RS3 retrograded starch RS4chemically modified starch and RS5 amylose-lipid com-plexes Chemical modification has been known to not onlyraise in vitro digestion resistance of starches reducingpostprandial glucose and insulin concentration but alsomaintain sensory attributes of the final foods [7] Degra-dation of RS in the large intestine has physiological

HindawiJournal of ChemistryVolume 2019 Article ID 2903252 10 pageshttpsdoiorg10115520192903252

benefits including the stimulation of intestinal bacterialactivity by prebiotic effect leading to the microbial fer-mentation and production of short-chain fatty acids whicheventually decrease the pH in colon [6] Upadhyaya et al[8] had reported that consumption of RS4 led to anabundance of Bifidobacterium adolescentis Parabacteroidesdistasonis and Christensenella minuta in the gut In ad-dition RS also has health beneficial effects such asprevention of colon cancer hypoglycemic effects hypo-cholesterolemic effects and inhibition of fat accumulation[9 10]

Polyfunctional carboxylic acids such as malic tartariccitric and glutaric acid have been used in the synthesis andrheological characterization of hydrogels [11] Xie and Liu[12] used citric acid and high temperature to increase theRS content of corn starch Compared to inorganic acidscitric acid is nutritionally harmless and increasing thedegree of substitution (DS) of starch by ester bond with thecitrate decreased the rate of digestion by pancreatin [9 13]Kim et al [14] reported that glutaric acid treatment at115degC affected the structural and digestion properties ofadley starch but lost the Maltese cross pattern in itsgranule Studies on citric acid-rice starch by Shin et al [15]showed that acid treatment hydrolyzed the branchingpoints of amylopectin leading to an increased apparentamylose content According to Kim and Shin [9] astructural difference such as in the number of carboxylgroups can affect the physicochemical properties ofstarch However the cross-linking capability of malic acidin reference to starch has not yet been reported Malic acidis a C4 carboxylic acid with two carboxyl groups com-prising 69ndash92 of all organic acids in grape berries andleaves [16] It is also naturally produced by many or-ganisms without showing any nutritional harm e USFood and Drug Administration classifies malic acid asldquogenerally recognized as safe (GRAS)rdquo and Food Chem-icals Codex (FCC) specifications list DL-malic acid as afood-grade organic acid [17] In industries Malic acid hasbeen used as food additive in the United States and Eu-ropean Union

Sweet potato Ipomoea batatas is a creeping di-cotyledonous plant belonging to the Convolvulaceaefamily Sweet potato has been a traditional source of starchin Asian countries and is one of the worldrsquos most im-portant food crops used as an ingredient in variousproducts such as noodles breads and cakes [18] epotential supply of sweet potato starch is exhaustive andcost-efficient In addition its components such as dietaryfiber carotenoids vitamins and minerals are healthbeneficial [19] erefore industrial interest is highly fo-cused on the use of native and modified sweet potatostarch Consequently extensive research regarding thechemicalphysicalenzymatic modifications of sweet po-tato starch should be performed to deepen the un-derstanding of its functional properties

e purposes of this study were to produce sweet potatostarch with low digestion property and heat stability bymalicacid treatment as well as to assess its physicochemicalproperties maintaining granule shape

2 Materials and Methods

21 Materials Sweet potato starch was purchased fromSeoahn Co (Buan Jeollabuk-do Korea) and DL-malic acid(M1210) was from Sigma Aldrich (St Louis MO USA) eenzymes used in starch digestion were porcine pancreatin(P7545 activity 8 times USPg Sigma) and amyloglucosidase(AMG 300L activity 300 AGUmL Novozymes BagsvaerdDenmark)

22 Preparation ofMalic Acid-Treated Starch DL-malic acidwas dissolved in water to prepare solutions of variousconcentrations (05 10 15 and 20M) with pH adjusted to35 by 10M NaOH Sweet potato starch (20 g) and 20mL ofdifferent concentrations of malic acid solution were mixedand kept in a stainless-steel bowl for 16 h at room tem-perature e bowls were then placed in an air-drying ovenat 50degC for 24 h e dried mixture was ground and placedeither in an air-drying oven at 130degC or at room temperaturefor 12 h It was washed thoroughly with distilled water toremove unreacted malic acid dried in an air drying oven at50degC and ground Samples were named according to theirprocessing condition in the concentration-temperatureformat A starch sample which underwent the same pro-cedure with distilled water (DW-130) and with 20M malicacid solution at 25degC for 16 h (2M-25) was used as thecontrol

23 Optical Microscopy Malic acid-treated starches wereobserved under a light microscope (CSB-HP3 Sam WonScientific Seoul Korea) with and without a polarizing plateGlycerol was used to disperse the sample on a glass slide withminimal air bubbles A digital camera (Nikon Tokyo Japan)was used to take the photographs

24 Fourier Transform-Infrared Spectroscopy (FT-IR)FT-IR spectra (VERTEX80v Bruker Billerica MA USA)were used to obtain the IR spectra e spectra were mea-sured ranging from 4000 to 600 cmminus1 in the transmissionmode at a resolution of 4 cmminus1 and normalized using the1315 cmminus1 peak of starch CH2 vibrations e samples werediluted with KBr (1 100 vv) before acquisition

25 Degree of Substitution (DS) Degree of substitution wasdetermined to estimate the average number of hydroxylgroups substituted with malic acid per anhydroglucose unitin starch e measurement was performed following themethod of by Xu et al [20] with some modifications Malicacid-treated starch (05 g) was placed in a 250mL glassbeaker with 50mL of distilled water e pH was measuredafter stirring themix for 1 h at 30degC To each beaker 25mL of05N NaOH was added to release the substituted groupsfrom the malic acid-treated starch and the solution wasstirred for 24 h at 50degC e excess NaOH was titrated backto original pH with 005NHCl DS was calculated as follows

2 Journal of Chemistry

DS 162 times NNaOH times VNaOH minus NHCl times VHCl( 1113857

1 000 times W minus 11609 times NNaOH times VNaOH minus NHCl times VHCl( 1113857 (1)

where DS is the degree of substitution W is the sampleweight (g) NNaOH is the normality of NaOH VNaOH is thevolume of NaOH NHCl is the normality of HCl 161 isglucose molecular weight 11609 is malic acid molecularweight used to back titrate and VHCl is the volume of HClused for back titration

26 InVitroDigestibility In vitro digestibility was measuredfollowing the method of Englyst et al [1] with slightmodifications by Shin et al [15] To prepare enzyme solu-tions porcine pancreatin (2 g) was added to 24mL distilledwater in a 50mL glass beaker and stirred for 10min esolution was then centrifuged at 1500 times g for 10min at 4degCto obtain a cloudy supernatante supernatant (20mL) wasmixed with 04mL of amyloglucosidase and 36mL of dis-tilled water To a 2mL microtube with 30mg of starchsample 075mL of sodium acetate buffer (01M pH 52) anda glass bead were added and either cooked for 30min or notcooked at all After cooling the tube to 37degC 075mL of theenzyme solution was added and incubated in a shakingincubator (240 rpm) e tubes were taken out after 10 and240min boiled for 10min in a heating block to stop thereaction and cooled to room temperature e tubes werecentrifuged at 5000 times g for 10min at 4degC e amount ofglucose in the supernatant was measured by the GOD-PODkit (Embiel Co Gunpo Korea) e amount of glucose after10min of enzyme reaction at 37degC indicated RDS and thatobtained after incubation for 10ndash240min was SDS RS wasthe starch not hydrolyzed after 240min of incubation

27 X-Ray Diffraction X-ray diffraction analysis was per-formed using an X-ray diffractometer (D8 ADVANCE withDAVINCI Bruker Karlsruhe Germany) operating at 40 kVand 40mA producing CuKα radiation of 15418 A wave-length scanning through the 2θ range of 3minus30deg and having astep time of 05 sec e relative crystallinity was calculatedusing the software developed by the instrument manufac-turer (EVA 20)

28 gtermal Properties ermal properties were de-termined using a differential scanning calorimeter (PyrisDiamond DSC Perkin-Elmer Waltham MA USA) Dis-tilled water (40 microL) was added to 10mg of the sample in astainless-steel DSC pan and sealede pan was kept at roomtemperature for more than 4 h for equilibration and uniformmixing e sample pan was heated gradually from 30degC to130degC at 5degCmin with an empty pan as the reference Toavoid condensation during the scan dry nitrogen wasflushed in the space surrounding the sample chamber Onset(To) peak (Tp) and conclusion (Tc) temperatures as well asgelatinization enthalpies (ΔH) were measured using thePyris software

29 Apparent Amylose Content Apparent amylose contentwas measured according to the colorimetric method out-lined by the AACC International Approved Method 61ndash03[21] e samples (20mg) were precisely weighed in 15 mLtubes and dispersed with 200 microL of absolute ethanol etubes were boiled for 10min after adding 18mL of 50NaOH e cooled solution (1mL) was placed in each tubewith 9mL of distilled water Diluted sample solutions wereput into a 15mL tube containing 9mL of distilled water and100 microL of 1N acetic acid Lugol solution (200 microL 02I2 + 20 KI Sigma) was added and kept in dark for 20minAbsorbance of the colored sample solution was measured at620 nm

210 Pasting Properties A Rapid Visco Analyzer (RVA-3DNewport Scientific Warriewood Australia) was used toinvestigate the pasting properties of sweet potato starchand malic acid-treated starch For each analysis 25 g ofstarch was added to an RVA canister with 25mL of distilledwater e measurement followed the AACC standardmethod 2 which includes a 23min heating and coolingprofile

211 Statistical Analyses All experiments were triplicatedand the mean values and standard deviations are reportedDuncanrsquos multiple range test was used to analyze the var-iance and the mean separations (plt 005) e statisticalanalyses were conducted using SPSS for Windows 220software (IBM Armonk NY USA)

3 Results and Discussion

31 Photomicrographs e granular shape of the variousmalic acid-treated starch samples was investigated using alight microscope (Figure 1) e shape of starches includingthe malic acid-treated samples had round semioval ovalspherical and round polygonal shapes which was consistentwith preivious studies [19 22] e malic acid-treated starchgranules were not ruptured even at malic acid concentra-tions as high as 20M According to Hirashima et al [23]granules of starch were all broken at pH below 30 whenmore glucose chains were observed compared to those inhigher pH-treated samples However in the range of pH40ndash60 the granule shape was retained while at pH above35 less glucose chains leached out and less fracture ofstarch granules occurred [23] Beyond pH 35 the granularshape of the malic acid-treated starches was not ruptureddespite the high concentration of malic acid e Maltesecross was observed using a polarizing plate confirming theinner ordered semicrystalline structure and radially orderedalignment of amylose and amylopectin [14 24] All samplesshowed the Maltese cross implying that the regular orderedinner structure of starch was mostly retained even after thethermal treatment in the presence of malic aciderefore it

Journal of Chemistry 3

(a)

(b)

(c)

(d)

(e)

Figure 1 Continued


could be assumed that the changes observed were caused notby the change of granular shape but by the effect of malicacid on the inner structure of starch

32 FT-IR FT-IR spectroscopy can determine the structuralcharacteristics in terms of the functional groups in malicacid-treated starches e peak in the range of 3000 to3500 cmminus1 indicates the presence of hydroxyl groups instarch and the one at 2930 cmminus1 indicates C-H bondstretching [25] e peak between 980 and 1200 cmminus1 can beattributed to C-O stretching vibration [26] e peak near1730 cmminus1 is a carbonyl peak [14 27] and the starches thatreacted with malic acid at 130degC commonly had a re-markable carbonyl peak at 1722 cmminus1 regardless of malicacid concentration (Figure 2) However the starches kept atroom temperature for 12 h or at 130degC without malic acidhad no carbonyl peak ese results could be explained bythe destruction of some parts of inter- and intramolecularhydrogen bonds by heat thereby leading to the formation ofan ester bond between starch and malic acid e peakintensity of thermally treated samples with a higher con-centration of malic acid was higher compared with thosewith low concentration of malic acid as shown in Figure 2is intensity appeared to be highly correlated to bothdegrees of substitution and RS content of malic acid-treatedstarch As the peak intensity of samples increased thecontent of RS also gradually increased (plt 005) e RScontent of DW-130 and 20M-130 were 270 and 534respectively is result was consistent with the report onglutarate-treated starch by Kim et al [14]

(f )

(g)

Figure 1 Light micrographs of raw and malic acid-treated sweet potato starches (magnification 400times) (a) raw (b) 20M-25 (c) DW-130(d) 05M-130 (e) 10M-130 (f ) 15M-130 and (g) 20M-130 1-under visible light and 2-under polarized light

Raw

Wavelength (cmndash1)1000150020002500300035004000

20M-25

DW-130

05M-130

10M-130

15M-130

20M-130

Figure 2 FT-IR spectra of raw and malic acid-treated sweet potatostarches


33 Degree of Substitution (DS) e DS of thermally treatedsamples is shown in Table 1 e value increased with theconcentration of malic acid While 20M-130 had the highestDS value of 0214 05M-130 had the lowest DS (0088)among the malic acid-treated starches is substitutionlevel was much higher than that in citric acid-substitutedstarches (0027) reacted at 1284degC for 138 h [15] DS isgenerally related to the number of carboxyl groups in or-ganic acids and the steric hindrance between them whichcan interrupt the ester bond formation For example aceticacid which has only one carboxylic group and smallermolecular size compared to other organic acids has theability to reach a high substitution level [20] Malic acidwhich has two carboxyl groups can theoretically form twoester bonds whereas citric acid can make three ester bonds

34 In Vitro Digestion of Samples Table 1 presents theproportion of RDS SDS and RS fractions e concentra-tion gradient of malic acid without thermal treatmentshowed no significant difference in RS content (data notshown) Regarding the digestibility of DW-130 raw and20M-25 there were no significant differences between RSand SDS content but a difference was observed in RDSwhich could have been due to the high heat treatment Onthe other hand thermal treatment along with malic acidincreased the content of RS ermally and malic acid-treated starch showed an increased RS content from270 (DW-130) to 534 (20M-130)eDS of substitutedstarches and RS fractions were highly correlated (r 0967plt 001) In addition malic acid and heat treatment de-creased SDS from 518 (DW-130) to 472 (20M-130)depending on the concentration of the malic acid solutionHowever the content of RDS was the highest in 10M-130(485) and decreased with increasing concentrations ofmalic acid is result suggested that some parts of theamylose and amylopectin structure were destroyed underacidic heating conditions so the RDS fraction increased until10M-130 but the other part forming ester bonds with malicacid became the RS fraction According to Huber amp BeMiller[28] the more the cross-linked starch the more the entry ofα-amylase molecules through the starch porous channelinhibited and hence more resistance to digestion ere-fore as the DS of malic acid-treated starch increased in-trusion of α-amylase was inhibited by the ester bond

between malic acid and starch chain However if the crosslink fully blocked α-amylase intrusion there should be noincrease of RDS e interference of cross links in thecomplexation of α-amylase and starch might be anotherreason for the increased RS [9 29] Despite the diffusion ofα-amylase after granule intrusion the cross-linked part ofstarch chain resists digestion

Since the starch used in food industry usually undergoesa cooking step high heat treatment could possibly destroythe ester bond thereby decreasing RS Digestion fractions ofcooked samples were also investigated Apart from thenonreacted starches that showed considerable decrease in RScontent 20M-130 had 499 of RS fraction which reducedfrom 534 Other malic acid-treated starches also haddecreased RS content but they were still highly correlatedwith the DS (r 0983 plt 001) e proportion of the RSfraction of DW-130 samples decreased from 270 to 195upon cooking e cooking procedure changed the RSproportions drastically from that of the nonthermallytreated samples whereas the thermally malic acid-treatedsamples had high content of heat-stable RS in high amountwhich indicates the presence of a rigid ester bond

35 X-Ray Diffraction e X-ray diffraction patterns areshown in Figure 3 e internal order of a starch granule isdemonstrated by X-ray diffraction patterns of A B and Ctypes [30] Native sweet potato starch showed the C-typepattern with diffraction intensities at 56deg 115deg 153deg 174deg232deg and 263deg at the angle 2θ C-type starches have beensubclassified into Ca Cb and Cc on the basis of their re-semblance to either A-type B-type or that between A-typeand B-type respectively [31] and the sweet potato starchused in this study belonged to the Cb-type Malic acid-treated starches maintained the same X-ray diffracto-grams but with differences in crystallinity and peak in-tensity As the concentration of malic acid solutionincreased the peak intensity and crystallinity decreased(DW-130 279 2M-130 176) indicating the influenceof heat-moisture treatment and the possibility of acid hy-drolysis e decreased crystallinity of DW-130 was seemedto be caused by the heat-moisture treatment Lee et al [32]reported that hydrothermal treatment decreased the majorpeaksrsquo intensities and their crystallinity e crystallinity ofmalic acid-treated samples was lower than that of raw starch

Table 1 Degree of substitution and in vitro digestibility of raw and malic acid-treated sweet potato starches

Sample DSNoncooked starch Cooked starch

RDS () SDS () RS () RDS () SDS () RS ()Raw nd 145plusmn 120a 529plusmn 420d 326plusmn 307bc 596plusmn 590c 198plusmn 245d 206plusmn 352a20M-25 nd 155plusmn 025a 552plusmn 241d 293plusmn 224ab 679plusmn 338de 137plusmn 404c 184plusmn 080aDW-130 nd 211plusmn 073b 518plusmn 176d 270plusmn 179a 718plusmn 371e 867plusmn 365b 195plusmn 116a05M-130 0088plusmn 00068a 364plusmn 153c 287plusmn 119c 349plusmn 151c 699plusmn 179e 357plusmn 265ab 266plusmn 427b10M-130 0127plusmn 00094b 485plusmn 273e 128plusmn 087b 387plusmn 186d 640plusmn 031cd 183plusmn 069a 342plusmn 096c15M-130 0184plusmn 00058c 469plusmn 116e 663plusmn 186a 465plusmn 283e 528plusmn 155b 387plusmn 261ab 433plusmn 193d20M-130 0214plusmn 00029d 419plusmn 074d 472plusmn 070a 534plusmn 069f 452plusmn 084a 486plusmn 239ab 499plusmn 257e

DS degree of substitution RDS rapidly digestible starch SDS slowly digestible starch RS resistant starch nd not detected e values with differentsuperscripts in each column are significantly different (plt 005) by Duncanrsquos multiple range test


due to acid hydrolysis and heat treatment during prepara-tion is result was consistent with the report on glutarate-treated starch by Kim et al [14] and citrate-treated starch byXie et al [33] Hydrogen bonds are known to sustain adouble helical structure but when substituted by esterbonds changes in double helical structure induce rear-rangement of the crystalline and semicrystalline structurehence lowering relative crystallinity

36 Gelatinization Parameters e gelatinization parame-ters of various malic acid-treated starches are shown inTable 2 Raw and 20M-25 samples had no signicant dif-ference in the To Tp Tc ΔH and even TcminusTo However DW-130 had lower To (577) Tp (666) Tc (713) and TcminusTo (149)and higher ΔH (136) which could have resulted from theheat-moisture treatment Increasing the concentration ofmalic acid decreased To Tp Tc and ΔH but increased TcminusToConsequently 20M-130 had lower To (440) Tp (508) Tc(620) and ΔH (289) and higher TcminusTo (181) compared withother samples DSC measures the primary hydrogen bondsthat stabilize the double helices within the granules [34] andthe quality and quantity of crystalline area by measuring the

change in heat energy [30]e decreased To Tp Tc ΔH andincreased TcminusTo in this study suggested that the internalcrystalline structure and helical structure of the malic acid-treated starch could have been disrupted and become aheterogeneous structure with rearrangement compared tothe unmodied starches If most of the crystalline area wasdestroyed no peak would be expected in the X-ray dif-fractogram however the X-ray diraction patterns of malicacid-treated starch were almost the same as those of rawstarch and nonthermally treated starches erefore malicacid which penetrated into starch granules may have notonly partially hydrolyzed the starch chain into shorterchains but also rearranged the crystalline structure of thegranules by substituting the hydrogen bonds with esterbonds With increased short chain melting temperaturedecreased which could be highly related to the decrease ofSDS and increase of RDS Decrease of ΔH corresponds to areduced amount of hydrogen bonds and is related to in-creased DS and a higher fraction of heat-stable RS

37 Apparent Amylose Content e apparent amylosecontents of various malic acid-treated starches are presentedin Table 3 Compared to the raw and nonthermal groupsamples with high DS values showed decreased apparentamylose content Apparent amylose contents of 20M-25 andDW-130 showed no signicant dierence with raw starch(pgt 005) However the increasing concentration of malicacid with heating decreased the apparent amylose content ofthe samples and that of 20M-130 was the lowest (207)e citric acid-treated starch had increased amount of ap-parent amylose content with the same treatment because ofthe hydrolysis which mainly occurred at the branchingpoint [15] but the opposite trend was observed in this studyMussulman and Wagoner [35] and Robin et al [36] sug-gested that acid hydrolysis mainly occurred during theconditioning step However structural analyses showed thatmost of the starch chain breakdown occurred during theheating step under the inuence of both acid and heatConsequently the decline of apparent amylose content wasdue to the rearrangement of the starch helix by the esterbond upon malic acid treatment and there could be in-creased amount of short chains due to hydrolysis which aretoo short to form the iodine complex

38 Pasting Properties e pasting properties of raw andmalic acid-treated starches are shown in Figure 4 e use ofthermal treatment with malic acid can lead to lower peakviscosity because of less swollen granules Cross-linkedstarches reacted under low pH conditions are reported tohave reduced swelling power [37] Starch samples whichwere only conditioned in malic acid solution (20M-25)showed a similar RVA curve with raw starch Raw starch and20M-25 had peak viscosity at 883degC and also had similarsetback viscosity As DW-130 indicated changes in structurewhich seemed to be the eect of slight the heat-moisturetreatment lower peak viscosity and setback viscosity weredetected by RVA which could also be due to the eect ofheat-moisture treatment Malic acid-treated starches had

Raw(306)

2θ5 10 15 20 25 30

20M-25(255)

DW-130(279)

05M-130(256)

10M-130(233)

15M-130(179)

20M-130(176)

Figure 3 X-ray diraction patterns and relative crystallinity of rawand malic acid-treated sweet potato starches Numbers in paren-theses indicate the percentage of crystallinity


lower and linear RVA proles (10M-130 15M-130 and20M-130) is result was similar to the dramatically de-creased pasting viscosity and linear RVA curve of citric acid-treated starch due to its nonswelling property [12] ehigher the concentration of malic acid solution used thelesser the viscosity observed Shukri and Shi [38] had re-ported that high level of cross-linking inhibits the swelling ofstarch granules because of less hydrogen bonding betweenthe helical structures in starch Similar to the previousstudies on citrate and acetate starches that reported in-creased stability of cooked starch as compared to nativestarch [37] malic acid-treated starch with high DS alsoshowed heat-stability not only toward gelatinization but alsotoward retrogradation due to the malic acid cross link

4 Conclusions

Heat treatment with a high concentration of malic acidsolution on sweet potato starch caused considerablechanges in the internal structure of starch maintaining itsgranular shape Moreover DS values and FT-IR spectrashowed ester bond formation between malic acid and thestarch e RS fraction of sweet potato starch drasticallyincreased with the ester bond formation and these RSfractions had remarkable heat-stable characteristicsStructural analyses by light microscopy XRD DSC RVAand AAC obviously demonstrated a low degree of hy-drolysis in starch chains (even at pH 35 of malic acidsolution) upon thermal treatment with malic acid andrearrangment of the crystalline area and alterations of thedouble helix by the substitution of the hydrogen bond byan ester bond Rapid visco analyzer (RVA) showed nopasting property of malic acid-treated starches due to itsnonswell properties caused by the malic acid cross-linkis information about the structural characteristics andheat-stable properties of RS in digestion can be used todevelop a low-digestible food ingredient and lead tofurther application of the study

Data Availability

e data used to support the ndings of this study areavailable from the corresponding author upon request

Disclosure

is article is based on the thesis submitted by ChinwooKwon as part of the requirements for his masterrsquos degree atSeoul National University [39]

Conflicts of Interest

e authors declare that they have no conicts of interestregarding the publication of this article

Acknowledgments

is work was supported by the National Research Foun-dation of Korea (NRF) grant funded by the Korean gov-ernment (MSIP) (no NRF-2016R1C1B2015506)

Table 2 Gelatinization parameters of raw control and malic acid-treated starches

Sample To (degC) Tp (degC) Tc (degC) TcminusTo (degC) ∆H (Jg)Raw 605plusmn 028e 682plusmn 007f 724plusmn 159d 119plusmn 183a 161plusmn 089d20M-25 607plusmn 021e 682plusmn 016f 726plusmn 037d 118plusmn 031a 166plusmn 046dDW-130 577plusmn 013d 666plusmn 031e 713plusmn 064d 136plusmn 077ab 149plusmn 066d05M-130 524plusmn 019c 639plusmn 025d 684plusmn 026c 160plusmn 033bc 121plusmn 163c10M-130 460plusmn 010b 601plusmn 048c 653plusmn 036b 194plusmn 026c 105plusmn 154c15M-130 454plusmn 073ab 530plusmn 047b 635plusmn 019ab 182plusmn 073c 750plusmn 107b20M-130 440plusmn 204a 508plusmn 102a 620plusmn 246a 181plusmn 434c 290plusmn 072a

To onset temperature Tp peak temperature Tc conclusion temperature TcminusTo temperature range of crystal melting ΔH enthalpy change e values withdierent superscripts in each column are signicantly dierent (plt 005) by Duncanrsquos multiple range test

Table 3 Apparent amylose content of raw and malic acid-treatedsweet potato starches

Sample Apparent amylose content ()Raw 269plusmn 075d20M-25 264plusmn 059cdDW-130 264plusmn 013cd05M-130 254plusmn 063c10M-130 230plusmn 068b15M-130 236plusmn 065b20M-130 207plusmn 042a

e values with dierent superscripts in each column are signicantlydierent (plt 005) by Duncanrsquos multiple range test

40

50

60

70

80

90

100

0

1000

2000

3000

4000

5000

0 200 400 600 800 1000 1200 1400

Tem

pera

ture

(degC)

Visc

osity

(cP)

Time (sec)

Figure 4 Rapid visco analyzer raw and malic acid-treated sweetpotato starches raw starch 20M-25 DW-130 05M-130 10M-130 15M-130 20M-130 temperature


References

[1] H N Englyst S M Kingman and J H Cummings ldquoClas-sification and measurement of nutritionally important starchfractionsrdquo European Journal of Clinical Nutrition vol 46no 2 pp S33ndashS50 1992

[2] I Gontildei L Garcıa-Diz E Mantildeas and F Saura-CalixtoldquoAnalysis of resistant starch a method for foods and foodproductsrdquo Food Chemistry vol 56 no 4 pp 445ndash4491996

[3] D Sievert and Y Pomeranz ldquoEnzyme-resistant Starch 1Characterization and evaluation by enzymatic thermoana-lytical and microscopic methodsrdquo Cereal Chemistry vol 66pp 342ndash347 1989

[4] J Tovar Bioavailability of starch in processed legumes PhDthesis Department of Applied Nutrition and Food ChemistryUniversity of Lund Sweden Lund Sweden 1992

[5] S G Haralampu ldquoResistant starch-a review of the physicalproperties and biological impact of RS3rdquo CarbohydratePolymers vol 41 no 3 pp 285ndash292 2000

[6] A P Nugent ldquoHealth properties of resistant starchrdquoNutritionBulletin vol 30 no 1 pp 27ndash54 2005

[7] M Stewart and J Zimmer ldquoA high fiber cookie made withresistant starch type 4 reduces post-prandial glucose andinsulin responses in healthy adultsrdquo Nutrients vol 9 no 3p 237 2017

[8] B Upadhyaya L McCormack A R Fardin-Kia et al ldquoImpactof dietary resistant starch type 4 on human gut microbiota andimmunometabolic functionsrdquo Scientific Reports vol 6 no 1p 28797 2016

[9] W Kim and M Shin ldquoEffects of organic acids and starchwater ratios on the properties of retrograded maize starchesrdquoFood Science and Biotechnology vol 20 no 4 pp 1013ndash10192011

[10] M G Sajilata R S Singhal and P R Kulkarni ldquoResistantstarch-A reviewrdquo Comprehensive Reviews in Food Science andFood Safety vol 5 no 1 pp 1ndash17 2006

[11] C Seidel W-M Kulicke C Heszlig B HartmannM D Lechner and W Lazik ldquoInfluence of the cross-linkingagent on the gel structure of starch derivativesrdquo Starch-Starkevol 53 no 7 pp 305ndash310 2001

[12] X Xie and Q Liu ldquoDevelopment and physicochemicalcharacterization of new resistant citrate starch from differentcorn starchesrdquo Starch-Starke vol 56 no 8 pp 364ndash3702004

[13] H Klaushofer E Berghofer and W Steyrer ldquoStarkecitrate -produktion und anwendungs-technische EigenschaftenrdquoStarch-Starke vol 30 no 2 pp 47ndash51 1978

[14] M J Kim S J Choi S I Shin et al ldquoResistant glutarate starchfrom adlay preparation and propertiesrdquo CarbohydratePolymers vol 74 no 4 pp 787ndash796 2008

[15] S I Shin C J Lee D-I Kim et al ldquoFormation character-ization and glucose response in mice to rice starch with lowdigestibility produced by citric acid treatmentrdquo Journal ofCereal Science vol 45 no 1 pp 24ndash33 2007

[16] A N Lakso andWM Kliewer ldquoe Influence of temperatureon malic acid metabolism in grape berries I enzyme re-sponsesrdquo Plant Physiology vol 56 no 3 pp 370ndash372 1975

[17] I Goldberg J S Rokem and O Pines ldquoOrganic acids oldmetabolites new themesrdquo Journal of Chemical Technology ampBiotechnology vol 81 no 10 pp 1601ndash1611 2006

[18] S I Shin H J Kim H J Ha S H Lee and T W MoonldquoEffect of hydrothermal treatment on formation and struc-tural characteristics of slowly digestible non-pasted granular

sweet potato starchrdquo Starch - Starke vol 57 no 9pp 421ndash430 2005

[19] F Zhu and S Wang ldquoPhysicochemical properties molecularstructure and uses of sweetpotato starchrdquo Trends in FoodScience amp Technology vol 36 no 2 pp 68ndash78 2014

[20] Y Xu V Miladinov and M A Hanna ldquoSynthesis andcharacterization of starch acetates with high substitutionrdquoCereal Chemistry Journal vol 81 no 6 pp 735ndash740 2004

[21] American Association of Cereal Chemists Approved Methodsof the AACC International Methods 61-03 AACC In-ternational St Paul MN USA 10th edition 2000

[22] S J Tian J E Rickard and J M V Blanshard ldquoPhysico-chemical properties of sweet potato starchrdquo Journal of theScience of Food and Agriculture vol 57 no 4 pp 459ndash4911991

[23] M Hirashima R Takahashi and K Nishinari ldquoEffects ofcitric acid on the viscoelasticity of cornstarch pastesrdquo Journalof Agricultural and Food Chemistry vol 52 no 10pp 2929ndash2933 2004

[24] W Blaszczak J Fornal S Valverde and L GarridoldquoPressure-induced changes in the structure of corn starcheswith different amylose contentrdquo Carbohydrate Polymersvol 61 no 2 pp 132ndash140 2005

[25] A Bajpai and J Shrivastava ldquoIn vitro enzymatic degradationkinetics of polymeric blends of crosslinked starch and car-boxymethyl celluloserdquo Polymer International vol 54 no 11pp 1524ndash1536 2005

[26] A Zarski S Ptak P Siemion and J Kapusniak ldquoEsterifi-cation of potato starch by a biocatalysed reaction in an ionicliquidrdquo Carbohydrate Polymers vol 137 pp 657ndash663 2016

[27] C N Saikia F Ali T Goswami and A C Ghosh ldquoEsteri-fication of high α-cellulose extracted from Hibiscus canna-binus Lrdquo Industrial Crops and Products vol 4 no 4pp 233ndash239 1995

[28] K C Huber and J N BeMiller ldquoChannels of maize andsorghum starch granulesrdquo Carbohydrate Polymers vol 41no 3 pp 269ndash276 2000

[29] K S Woo and P A Seib ldquoCross-linked resistant starchpreparation and propertiesrdquoCereal Chemistry Journal vol 79no 6 pp 819ndash825 2002

[30] D Cooke and M J Gidley ldquoLoss of crystalline and molecularorder during starch gelatinisation origin of the enthalpictransitionrdquo Carbohydrate Research vol 227 pp 103ndash1121992

[31] S Hizukuri M Fujii and Z Nikuni ldquoe effect of inorganicions on the crystallization of amylodextrinrdquo Biochimica etBiophysica Acta vol 40 pp 346ndash348 1960

[32] C J Lee S I Shin Y Kim H J Choi and T W MoonldquoStructural characteristics and glucose response in mice ofpotato starch modified by hydrothermal treatmentsrdquo Car-bohydrate Polymers vol 83 no 4 pp 1879ndash1886 2011

[33] X Xie Q Liu and S W Cui ldquoStudies on the granularstructure of resistant starches (type 4) from normal highamylose and waxy corn starch citratesrdquo Food Research In-ternational vol 39 no 3 pp 332ndash341 2006

[34] C G Biliaderis ldquoermal analysis of food carbohydratesrdquo ingtermal Analysis of Foods V R Harwalkar and C Y MaEds pp 168ndash220 Elsevier Science Publishers Ltd LondonUK 1990

[35] W Mussulman and J Wagoner ldquoElectron microscopy ofunmodified and acid-modified corn starchesrdquo CerealChemistry vol 45 pp 162ndash171 1968

[36] J P Robin C Mercier R Charbonniere and A GuilbotldquoLintnerized starches Gel-filtration and enzymatic studies of


insoluble residues from prolonged acid treatment of potatostarchrdquo Cereal Chemistry vol 51 pp 389ndash406 1974

[37] S O Agboola J O Akingbala and G B OguntimeinldquoPhysicochemical and functional properties of low DS cassavastarch acetates and citratesrdquo Starch-Starke vol 43 no 2pp 62ndash66 1991

[38] R Shukri and Y-C Shi ldquoStructure and pasting properties ofalkaline-treated phosphorylated cross-linked waxy maizestarchesrdquo Food Chemistry vol 214 pp 90ndash95 2017

[39] C Kwon ldquoStructural and physicochemical characteristics ofgranular malic acid-treated heat-stable sweet potato starchrdquoMD thesis Department of Agricultural Biotechnology SeoulNational University Seoul South Korea 2018


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Submit your manuscripts atwwwhindawicom

benefits including the stimulation of intestinal bacterialactivity by prebiotic effect leading to the microbial fer-mentation and production of short-chain fatty acids whicheventually decrease the pH in colon [6] Upadhyaya et al[8] had reported that consumption of RS4 led to anabundance of Bifidobacterium adolescentis Parabacteroidesdistasonis and Christensenella minuta in the gut In ad-dition RS also has health beneficial effects such asprevention of colon cancer hypoglycemic effects hypo-cholesterolemic effects and inhibition of fat accumulation[9 10]

Polyfunctional carboxylic acids such as malic tartariccitric and glutaric acid have been used in the synthesis andrheological characterization of hydrogels [11] Xie and Liu[12] used citric acid and high temperature to increase theRS content of corn starch Compared to inorganic acidscitric acid is nutritionally harmless and increasing thedegree of substitution (DS) of starch by ester bond with thecitrate decreased the rate of digestion by pancreatin [9 13]Kim et al [14] reported that glutaric acid treatment at115degC affected the structural and digestion properties ofadley starch but lost the Maltese cross pattern in itsgranule Studies on citric acid-rice starch by Shin et al [15]showed that acid treatment hydrolyzed the branchingpoints of amylopectin leading to an increased apparentamylose content According to Kim and Shin [9] astructural difference such as in the number of carboxylgroups can affect the physicochemical properties ofstarch However the cross-linking capability of malic acidin reference to starch has not yet been reported Malic acidis a C4 carboxylic acid with two carboxyl groups com-prising 69ndash92 of all organic acids in grape berries andleaves [16] It is also naturally produced by many or-ganisms without showing any nutritional harm e USFood and Drug Administration classifies malic acid asldquogenerally recognized as safe (GRAS)rdquo and Food Chem-icals Codex (FCC) specifications list DL-malic acid as afood-grade organic acid [17] In industries Malic acid hasbeen used as food additive in the United States and Eu-ropean Union

Sweet potato Ipomoea batatas is a creeping di-cotyledonous plant belonging to the Convolvulaceaefamily Sweet potato has been a traditional source of starchin Asian countries and is one of the worldrsquos most im-portant food crops used as an ingredient in variousproducts such as noodles breads and cakes [18] epotential supply of sweet potato starch is exhaustive andcost-efficient In addition its components such as dietaryfiber carotenoids vitamins and minerals are healthbeneficial [19] erefore industrial interest is highly fo-cused on the use of native and modified sweet potatostarch Consequently extensive research regarding thechemicalphysicalenzymatic modifications of sweet po-tato starch should be performed to deepen the un-derstanding of its functional properties

e purposes of this study were to produce sweet potatostarch with low digestion property and heat stability bymalicacid treatment as well as to assess its physicochemicalproperties maintaining granule shape

2 Materials and Methods

21 Materials Sweet potato starch was purchased fromSeoahn Co (Buan Jeollabuk-do Korea) and DL-malic acid(M1210) was from Sigma Aldrich (St Louis MO USA) eenzymes used in starch digestion were porcine pancreatin(P7545 activity 8 times USPg Sigma) and amyloglucosidase(AMG 300L activity 300 AGUmL Novozymes BagsvaerdDenmark)

22 Preparation ofMalic Acid-Treated Starch DL-malic acidwas dissolved in water to prepare solutions of variousconcentrations (05 10 15 and 20M) with pH adjusted to35 by 10M NaOH Sweet potato starch (20 g) and 20mL ofdifferent concentrations of malic acid solution were mixedand kept in a stainless-steel bowl for 16 h at room tem-perature e bowls were then placed in an air-drying ovenat 50degC for 24 h e dried mixture was ground and placedeither in an air-drying oven at 130degC or at room temperaturefor 12 h It was washed thoroughly with distilled water toremove unreacted malic acid dried in an air drying oven at50degC and ground Samples were named according to theirprocessing condition in the concentration-temperatureformat A starch sample which underwent the same pro-cedure with distilled water (DW-130) and with 20M malicacid solution at 25degC for 16 h (2M-25) was used as thecontrol

23 Optical Microscopy Malic acid-treated starches wereobserved under a light microscope (CSB-HP3 Sam WonScientific Seoul Korea) with and without a polarizing plateGlycerol was used to disperse the sample on a glass slide withminimal air bubbles A digital camera (Nikon Tokyo Japan)was used to take the photographs

24 Fourier Transform-Infrared Spectroscopy (FT-IR)FT-IR spectra (VERTEX80v Bruker Billerica MA USA)were used to obtain the IR spectra e spectra were mea-sured ranging from 4000 to 600 cmminus1 in the transmissionmode at a resolution of 4 cmminus1 and normalized using the1315 cmminus1 peak of starch CH2 vibrations e samples werediluted with KBr (1 100 vv) before acquisition

25 Degree of Substitution (DS) Degree of substitution wasdetermined to estimate the average number of hydroxylgroups substituted with malic acid per anhydroglucose unitin starch e measurement was performed following themethod of by Xu et al [20] with some modifications Malicacid-treated starch (05 g) was placed in a 250mL glassbeaker with 50mL of distilled water e pH was measuredafter stirring themix for 1 h at 30degC To each beaker 25mL of05N NaOH was added to release the substituted groupsfrom the malic acid-treated starch and the solution wasstirred for 24 h at 50degC e excess NaOH was titrated backto original pH with 005NHCl DS was calculated as follows














(a)

(b)

(c)

(d)

(e)

Figure 1 Continued




(f )

(g)


Raw


20M-25

DW-130

05M-130

10M-130

15M-130

20M-130


















Raw(306)

2θ5 10 15 20 25 30

20M-25(255)

DW-130(279)

05M-130(256)

10M-130(233)

15M-130(179)

20M-130(176)




4 Conclusions


Data Availability


Disclosure




Acknowledgments








40

50

60

70

80

90

100

0

1000

2000

3000

4000

5000

0 200 400 600 800 1000 1200 1400

Tem

pera

ture

(degC)

Visc

osity

(cP)

Time (sec)



References

















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls
















(a)

(b)

(c)

(d)

(e)

Figure 1 Continued




(f )

(g)


Raw


20M-25

DW-130

05M-130

10M-130

15M-130

20M-130


















Raw(306)

2θ5 10 15 20 25 30

20M-25(255)

DW-130(279)

05M-130(256)

10M-130(233)

15M-130(179)

20M-130(176)




4 Conclusions


Data Availability


Disclosure




Acknowledgments








40

50

60

70

80

90

100

0

1000

2000

3000

4000

5000

0 200 400 600 800 1000 1200 1400

Tem

pera

ture

(degC)

Visc

osity

(cP)

Time (sec)



References

















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls






(f )

(g)


Raw


20M-25

DW-130

05M-130

10M-130

15M-130

20M-130


















Raw(306)

2θ5 10 15 20 25 30

20M-25(255)

DW-130(279)

05M-130(256)

10M-130(233)

15M-130(179)

20M-130(176)




4 Conclusions


Data Availability


Disclosure




Acknowledgments








40

50

60

70

80

90

100

0

1000

2000

3000

4000

5000

0 200 400 600 800 1000 1200 1400

Tem

pera

ture

(degC)

Visc

osity

(cP)

Time (sec)



References

















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls



















Raw(306)

2θ5 10 15 20 25 30

20M-25(255)

DW-130(279)

05M-130(256)

10M-130(233)

15M-130(179)

20M-130(176)




4 Conclusions


Data Availability


Disclosure




Acknowledgments








40

50

60

70

80

90

100

0

1000

2000

3000

4000

5000

0 200 400 600 800 1000 1200 1400

Tem

pera

ture

(degC)

Visc

osity

(cP)

Time (sec)



References

















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls





4 Conclusions


Data Availability


Disclosure




Acknowledgments








40

50

60

70

80

90

100

0

1000

2000

3000

4000

5000

0 200 400 600 800 1000 1200 1400

Tem

pera

ture

(degC)

Visc

osity

(cP)

Time (sec)



References

















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls




References

















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls














Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls




structuralandphysicochemicalcharacteristicsofgranularmalic … · 2018. 9. 20. · ds 162 ×n naoh...

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