Effects of Amylopectin Brnch Chin Length nd Amylose Content on the Geltiniztion nd Psting Properties of Strch 1 J. Jne, 2,3 Y. Y. Chen, 2 L. F. Lee, 2 A. E. McPherson, 2 K. S. Wong, 2 M. Rdosvljevic, 2 nd T. Ksemsuwn 2 ABSTRACT Cerel Chem. 76(5):629 637 Structures nd properties of strches isolted from different otnicl sources were investigted. Apprent nd solute mylose contents of strches were determined y mesuring the iodine ffinity of deftted whole strch nd of frctionted nd purified mylopectin. Brnch chinlength distriutions of mylopectins were nlyzed quntittively using high-performnce nion-exchnge chromtogrphy system equipped with postcolumn enzyme rector nd pulsed mperometric detector. Therml nd psting properties were mesured using differentil scnning clorimetry nd rpid visconlyzer, respectively. Asolute mylose contents of most of the strches studied were lower thn their pprent mylose contents. This difference correlted with the numer of very long rnch chins of mylopectin. Studies of mylopectin structures showed tht ech strch hd distinct rnch chin-length distriution profile. Averge degrees of polymeriztion (dp) of mylopectin rnch chin length rnged from 18.8 for wxy rice to 30.7 for high-mylose mize VII. Compred with X-ry A-type strches, B-type strches hd longer chins. A shoulder of dp 18 21 (chin length of 6.3 7.4 nm) ws found in mny strches; the chin length of 6.3 7.4 nm ws in the proximity of the length of the mylopectin crystlline region. Strches with short verge mylopectin rnch chin lengths (e.g., wxy rice nd sweet rice strch), with lrge proportions of short rnch chins (dp 11 16) reltive to the shoulder of dp 18 21 (e.g., whet nd rley strch), nd with high strch phosphte monoester content (e.g., potto strch) displyed low geltiniztion tempertures. Amylose contents nd mylopectin rnch chin-length distriutions predominntly ffected the psting properties of strch. Strch consists of two polyscchrides, mylose nd mylopectin. Amylose hs long liner chins of (1 4)-linked α-d-glucopyrnose residues, some with few (>10) rnches (Hizukuri et l 1981). Amylopectin hs lrge moleculr weight nd highly rnched structures consisting of much shorter chins of (1 4) α-dglucose residues. The rnch-chins re connected y (1 6)-α-Dglucosidic linkges (Hizukuri 1986). Some strches lso contin third component known s the intermedite component (Lnsky et l 1949). Most strches contin 20 30% mylose nd 70 80% mylopectin; rtios vry with the otnicl source of the strch. The mylose-to-mylopectin rtio of strch gretly ffects the strch functionl properties. However, mylose contents of strches reported in literture re mostly pprent mylose (Morrison nd Lignelet 1983, Knutson nd Grove 1994). Tked nd Hizukuri (1987) hve reported tht strches isolted from indic rices hve more pprent mylose contents thn do jponic rices. However, indic rice mylopectins hve longer chins thn jponic rice mylopectins. Becuse long chins of mylopectin lso cn form helicl complex with iodine, the iodine ffinity nd mylose content of the strch re inflted. The solute (rel) mylose contents of the indic nd jponic rice strches re found to e similr. These results show tht mylopectin with long rnch chins cuse overestimtion of the mylose content in strch when it is determined y iodine titrtion or y the lue vlue method. The study of Ksemsuwn et l (1995) confirms tht long rnch chins of mylopectin interct with iodine nd result in higher iodine ffinity of the strch. Amylopectin is the mjor component of most strches, nd its fine structure plys criticl role in the chrcteristics of strch. Studies hve reveled tht the rnch chin-length of mylopectin is relted to the strch crystlline structure (Hizukuti et l 1983, Hizukuri 1985, Gidley nd Bulpin 1987, Hnshiro et l 1996). The rnch chin length lso ffects the geltiniztion, retrogrdtion (Miles et l 1985; Gudmundsson nd Elisson 1990; Klichevsky et l 1990; Jne et l 1992; Shi nd Sei 1992, 1995; Yun 1 Journl Pper No. J.-17972 of the Iow Agriculture nd Home Economics Experiment Sttion, Ames, IA. Project no. 3258. 2 Deprtment of Food Science nd Humn Nutrition nd Center for Crops Utiliztion Reserch, Iow Stte University, Ames, IA 50011. 3 Corresponding uthor. E-mil: jjne@istte.edu Puliction no. C-1999-0726-07R. 1999 Americn Assocition of Cerel Chemists, Inc. et l 1993; Lu et l 1997), nd psting properties of strch (Jne et l 1992, Jne nd Chen 1992, Wng et l 1993). The reltionship etween the mylopectin rnch chin length nd the functionl properties of strch isolted from different otnicl sources ws difficult to estlish ecuse different methods were used y different investigtors. Detiled informtion out the reltionship etween the structure nd functionl property of strch is needed to provide direction for genetic modifictions. In this study, the pprent nd solute mylose contents of strch were determined y mesuring the iodine ffinity of deftted whole strch nd of the frctionted pure mylopectin. The rnch chin-length distriution of mylopectin ws quntittively determined y using high-performnce nion-exchnge chromtogrphy system equipped with postcolumn enzyme rector nd pulsed mperometric detector (HPAEC-PAD-ENZ) (Wong nd Jne 1997). Therml properties (e.g., geltiniztion nd retrogrdtion) nd psting properties of the strch were studied y using differentil scnning clorimetry nd rpid visconlyzer, respectively. Results otined from these studies were used to estlish correltion etween chemicl structures nd functionl properties of strches. MATERIALS AND METHODS Mterils Chinese tro, mung en, wxy rice, sweet rice, green lef cnn, lotus root, wter chestnut, nd cttil millet strches were isolted in the lortory s reported y Lim et l (1994). Wxy mrnth I ws isolted y the lkline method (Myers nd Fox 1994). Wxy mrnth II nd norml glcier rley strch were isolted y comintion of mild lkline nd protese tretments (Rdosvljevic et l 1998). Green nn strch ws provided y A. R. Bonill, University of Cost Ric. Wxy mize, myloseextender (e) wxy nd dull (du) wxy mize strches were gifts of Cerestr, USA (Hmmond, IN); potto nd whet strches were purchsed from Sigm Chemicl Co. (St. Louis, MO); rice strch ws from Mtheson Colemn & Bel Inc. (Cincinnti, OH); lrge grnule rley strch ws gift of Oylko Co. (Finlnd); norml glcier rley ws gift of Wlter Newmn (Montn Stte University); nd tpioc, norml mize, nd high-mylose mize (V) nd (VII) strches were gifts of Ntionl Strch nd Chemicl Co. (Bridgewter, NJ). Isomylse (EC 3.2.1.68) from Pseudomons mylodermos ws purchsed from Hyshir Biochemicl Lortories, Inc. (Okym, Jpn), nd myloglucosidse (EC Vol. 76, No. 5, 1999 629
3.2.1.3) from Rhizopus mold (9032-80-0) ws purchsed from Sigm nd ws used without further purifiction. Methods Amylose nd mixture of mylopectin nd intermedite components were seprted following modified Schoch method (Jne nd Chen 1992). The mixture of mylopectin nd intermedite components ws purified y recrystllizing five times to remove mylose residues. The purity of mylopectin ws exmined y gel-permetion chromtogrphy monitored y the totl crohydrte nd lue vlue (Jne nd Chen 1992). Iodine ffinities of deftted strch nd of isolted mylopectin were mesured y using n utomtic potentiometric titrtor (702 SM Tirino, Metrohm, Herisu, Switzerlnd) following the method previously reported (Ksemsuwn et l 1995). Apprent mylose contents were clculted y dividing the iodine ffinity of deftted strch y 20% (Tked nd Hizukuri 1987). Asolute mylose contents were clculted y the method of Ksemsuwn et l (1995). Ech nlysis ws replicted minimum of three times. Amylopectin ws dernched y isomylse following the procedure of Jne nd Chen (1992). The mylopectin rnch chinlength distriutions were nlyzed y using high-performnce nion-exchnge chromtogrphy system equipped with n enzyme column rector nd pulsed mperometric detector (Dionex, Sunnyvle, CA) (HPAEC-ENZ-PAD) y the method reported y Wong nd Jne (1997). A DX-300 grdient chromtogrphy system, Cropc PA100 (4 250 mm) column, Cropc PA 100 gurd column (3 25 mm), nd n enzyme column (2 23 mm) immoilized with myloglucosidse were employed. The pulsed potentils nd durtions were E1 = 0.05 V (t1= 480 msec), E2 = 0.60 V (t2 =120 msec), nd E3 = -0.60 V (t3 = 60 msec). The eluents A nd B were 100 mm sodium hydroxide nd 100 mm sodium hydroxide in 300 mm sodium nitrte solution, respectively. The grdient of eluent B ws 1% t 0 min, 5% t 39 min, 8% t 50 min, 30% t 170 min, nd 45% t 220 min. The eluent grdient ws operted t 0.5 ml/min flow rte. The results were otined from t lest two replictes of ech mylopectin smple. Strch X-ry ptterns were otined with copper, nickel foilfiltered, K α rdition y using diffrctometer (D-500, Siemens, Mdison, WI). The diffrctometer ws operted t 27 ma nd 50 kv. The scnning region of the diffrction ngle (2θ) ws from 4 with 0.05 step size nd count time of 2 sec. Strch smples were equilirted in 100% rh chmer t 25 C for 24 hr efore mesurement. Geltiniztion nd retrogrdtion properties of strches were nlyzed y using differentil scnning clorimeter (DSC-7, Perkin- Elmer, Norwlk, CT) equipped with n intrcooling II system. Aluminum pns (Perkin-Elmer) were used for the nlyses. Strch smples ( 2 mg ech, ds) were weighed in smple pns, mixed with distilled wter ( 6 mg), nd seled. The heting rte ws 10 C/min over the temperture rnge of 25 100 C. Stinless steel pns (Perkin-Elmer) were used for high-mylose mize V nd VII strches t heting rnge of 25 150 C. Indium nd zinc were used s the reference stndrds. Enthlpy chnge ( H), geltiniztion onset temperture (T o ), pek temperture (T p ), nd geltiniztion rnges were computed utomticlly. The dt were verges of minimum of three replictes of ech strch smple. The retrogrdtion study ws performed using the sme method with the sme smple used for geltiniztion study fter storge t 4 C for seven dys. Strch psting properties were nlyzed y using Rpid ViscoAnlyser (RVA) (Newport Scientific, Sydney, Austrli). Ech strch suspension (8%, w/w, ds; 28 g of totl weight) ws equilirted t 50 C for 1 min, heted t rte of 6 C/min to 95 C, mintined t tht temperture for 5 min, nd then cooled to 50 C t rte of 6 C/min. A constnt rotting speed of the pddle (160 rpm) ws used. Becuse of the limited mount of smple, 5% (w/w, ds) of green lef cnn strch suspension ws used, nd it ws compred with potto nd norml mize strches t the sme concentrtion level. RESULTS AND DISCUSSION Asolute Amylose Content Long rnch chins of mylopectin, like mylose, ind iodine to form single helicl complex during potentiometric titrtion, develop lue color, nd, consequently, inflte the iodine ffinity nd the pprent mylose content of the strch. Becuse of this, we exmined oth the pprent mylose content nd the solute mylose content of ech strch, nd the results re shown in Tle I. Most norml strches, except Chinese tro, hd lower solute mylose contents thn their pprent mylose contents. Iodine ffin- TABLE I Iodine Affinities nd Amylose Contents of Strches Iodine Affinity Amylose Content (%) c Source Strch Amylopectin Apprent (A) d Asolute (B) e A B A-type strch Norml mize 5.88 ± 0.14 1.78 ± 0.03 29.4 22.5 6.9 Rice 5.00 ± 0.02 1.11 ± 0.00 25.0 20.5 4.5 Whet 5.75 ± 0.15 0.8 ± 0.2 28.8 25.8 3.0 Brley 5.1 ± 0.3 0.50 ± 0.04 25.5 23.6 1.9 Cttil millet 3.97 ± 0.06 1.08 ± 0.03 19.8 15.3 4.5 Mung en 7.58 ± 0.06 2.07 ± 0.03 37.9 30.7 7.2 Chinese tro 2.75 ± 0.06 0.00 ± 0.00 13.8 13.8 0.0 Tpioc 4.7 ± 0.2 1.39 ± 0.07 23.5 17.8 5.7 B-type strch Amylomize V 10.4 ± 0.1 6.79 ± 0.04 52.0 27.3 24.7 Amylomize VII 13.6 ± 0.2 9.3 ± 0.1 68.0 40.2 27.8 Potto 7.20 ± 0.01 4.6 ± 0.1 36.0 16.9 19.1 Green lef cnn 8.64 ± 0.00 5.31 ± 0.03 43.2 22.7 20.5 C-type strch Wter chestnut 5.79 ± 0.19 3.08 ± 0.02 29.0 16.0 13.0 Determined y X-ry diffrction. Averged from t lest three replictes ± stndrd devition of ech smple, except for deftted green lef cnn strch, of which the iodine ffinity ws verged from two replictes. c Iodine ffinity for pure mylose ws ssigned s 20% (Tked nd Hizukuri 1987). d Clculted s: C = 100 IA S /0.20 where C is the percentge of pprent mylose content nd IA S is the iodine ffinity of the whole deftted strch. e Clculted s: C = (IA S IA AP+IC )/{0.20 (IA AP+IC /100)} where C is the percentge of solute mylose content, IA S is the iodine ffinity of whole deftted strch, nd IA AP+IC is the iodine ffinity of the mylopectin nd the intermedite component mixture. 630 CEREAL CHEMISTRY
ities of wxy strches, which contined no mylose, re shown in Tle II. Among the wxy strches, e wxy mize strch displyed n pprent mylose content of 34.5%, despite the strch contining no mylose. Strches displying B-type X-ry pttern, which hd lrger proportions of long mylopectin rnch chins thn the A- nd C-type strches (Tle III), gve greter over-estimtions of the mylose content (Tles I nd II). The results of HPAEC-ENZ-PAD chromtogrphy (Tle III) showed tht mylopectin without detectle very long rnch chins (dp > 73) (Chinese tro mylopectin, wxy rice, nd wxy mize strches), displyed no detectle iodine ffinity (Tles I nd II). These results indicted positive correltion etween the iodine ffinity of mylopectin nd the presence of very long rnch chins of the mylopectin insted of long verge-rnch-chin lengths. Brnch Chin-Length Distriution of Amylopectin Normlized HPAEC-ENZ-PAD chromtogrms of rnch chinlength distriutions of mylopectins isolted from strches tht disply A-, B-, nd C-type X-ry ptterns re shown in Figs. 1 3, respectively, nd the computed results re summrized in Tle III. In generl, A-type strches hd peks t shorter chin-lengths TABLE II Iodine Affinities nd Apprent Amylose Contents of Wxy Strches Source Iodine Affinity Apprent Amylose Content c (%) A-type strch Wxy mize 0.00 ± 0.00 0.0 du Wxy mize 0.42 ± 0.03 2.1 Wxy rice 0.00 ± 0.00 0.0 Sweet rice 0.41 ± 0.03 2.1 Wxy mrnth 0.68 ± 0.03 3.4 B-type strch e Wxy mize 6.89 ± 0.19 34.5 Determined y X-ry diffrction. Iodine ffinities were verged from t lest three replictes ± stndrd devition. Iodine ffinity for pure mylose ws ssigned s 20% (Tked nd Hizukuri 1987). c Clculted s: C = 100 IA S /0.20 where C is the percentge of pprent mylose content nd IA S is the iodine ffinity of the whole deftted strch. (first pek t dp 12 14, second pek t dp 41 51) (Fig. 1) thn B- type strches (first pek t dp 14 16, second pek t dp 48 53) (Fig. 2). A-type strches lso hd lrger proportions (15.6 27.4%) of short chins (dp 6 12) nd smller proportions (6.6 26.7%) of long chins (dp > 37) thn B-type strches, 8.5 12.3% nd 26.1 29.5% for the short nd long chins, respectively (Tle III). The C-type strches hd sustntil mounts of oth short nd long rnch-chins (Fig. 3). Although some trends of mylopectin rnch chin-length distriutions were oserved for strches with the sme crystlline type, ech strch hd its own chin-length distriution profile (Figs. 1 3). For exmple, ll the cerel strches, nd mny other strches, displyed very few short chins of dp 6 nd grdul increse in chins of dp 7 9. Some tuer, root, nd legume strches such s Chinese tro, lotus root, wter chestnut, nd mung en strch displyed higher popultion of dp 7 thn dp 6 nd 8. Potto mylopectin hd the lowest popultion of dp 8. These results were in greement with those reported y Hnshiro et l (1996). The HPAEC-ENZ-PAD chromtogrms yielded more detiled results of the rnch chin-length distriution of mylopectins, especilly for the long rnch chins. Most strch smples nlyzed displyed shoulder t dp 18 21, which hs een reported y Hnshiro et l (1996). The reltive intensity of the shoulder differed with strches. Among the strches studied, whet, rley, du wxy mize, mrnth, nd tpioc hd the most ovious shoulders. Amylopectins of e wxy mize nd high-mylose mize V nd VII lso displyed shoulders; however, the reltive intensities of the shoulders were close to the pek height, much higher thn other strches (Fig. 2). Physicl distnces of strch chins with dp 18 21 re clculted to e 6.3 7.4 nm, using 0.35 nm for the distnce of ech glucose nhydrous unit. The rnge of the distnce, 6.3 7.4 nm, ws in the proximity of the length of mylopectin crystlline region, 6.65 nm for whet, reported y Cmeron nd Donld (1992). The results suggested tht dp 18 21 represented the full length of the crystlline region, nd the rtio of the intensities of pek 1 nd the shoulder indicted the proportion of short chins in the crystllites tht result in defects. Strches tht did not show shoulder in the chromtogrm include norml mize, norml rice, cttil millet, Chinese tro, potto, green-lef cnn, nd green nn. TABLE III Brnch Chin-Length (CL) Distriutions of Amylopectins Pek dp Averge % Distriution Highest Source I II CL dp 6 9 dp 6 12 dp 13 24 dp 25 36 dp 37 Detectle dp A-type strch Norml mize 13 48 24.4 3.85 17.9 47.9 14.9 19.3 80 Wxy mize 14 48 23.5 6.94 17.0 49.4 17.1 16.5 73 du Wxy mize 14 51 23.1 1.21 16.7 51.9 17.4 14.0 80 Norml rice 12 46 22.7 4.10 19.0 52.2 12.3 16.5 80 Wxy rice 12 41 18.8 8.57 27.4 53.4 12.6 6.6 66 Sweet rice 12 45 21.6 7.59 23.5 48.7 13.7 14.0 78 Whet 12 41 22.7 5.18 19.0 41.7 16.2 13.0 77 Brley 12 43 22.1 4.92 20.8 48.9 17.7 12.6 75 Wxy mrnth 12 43 21.8 6.31 25.1 47.7 12.5 14.7 80 Cttil millet 13 45 21.5 4.21 20.2 53.8 12.7 13.3 74 Mung en 13 48 24.8 2.45 15.6 47.6 18.3 18.5 74 Chinese tro 13 45 23.4 5.99 18.8 48.7 14.8 17.7 71 Tpioc 12 49 27.6 4.68 17.3 40.4 15.6 26.7 79 B-type strch e Wxy mize 16 53 29.5 2.29 10.4 43.5 18.1 28.0 84 Amylomize V 16 48 28.9 1.90 9.7 43.9 20.3 26.1 86 Amylomize VII 16 48 30.7 1.81 8.5 40.7 21.3 29.5 86 Potto 14 52 29.4 3.53 12.3 43.3 15.5 28.9 85 Green lef cnn 15 52 28.9 3.41 11.7 45.3 16.2 26.8 85 C-type strch Lotus root 13 52 25.4 4.57 16.4 47.2 15.4 21.0 83 Wter chestnut 13 50 26.7 5.86 17.8 43.7 15.3 23.2 80 Green nn 13 48 26.4 5.25 16.8 46.3 12.9 24.0 79 Grouping of degree of polymeriztion (dp) numers followed tht of Hnshiro et l (1996). Determined y X-ry diffrction. Vol. 76, No. 5, 1999 631
Fig. 1. Amylopectin rnch chin-length distriutions of A-type strches determined y using high-performnce nion-exchnge chromtogrphy system equipped with n enzyme column rector nd pulsed mperometric detector (HPAEC-ENZ-PAD). A Cropc PA100 column nd n immoilized myloglucosidse column were used for the nlysis. 632 CEREAL CHEMISTRY
Strch Geltiniztion Geltiniztion properties of strches mesured y differentil scnning clorimetry (DSC) re summrized in Tle IV. Among the strch smples, wxy rice, whet, rley, nd potto displyed the lowest onset geltiniztion tempertures. Wxy rice strch hd the shortest verge rnch chin length (dp 18.8), the shortest second pek chin length (dp 41), the lrgest proportion of short chins (dp 6 12, 27.4%), very few long chins, nd shoulder t dp 18. Studies with rice strches (Asok et l 1985), tro strch (Jne et l 1992), nd wxy mize strches (Shi nd Sei 1992, 1995, Yun et l 1993) hve shown tht those with shorter long-b chins disply lower geltiniztion tempertures. Whet nd rley strches, with verge chin lengths of dp 22.1 22.7 nd reltively short second pek chin lengths (dp 41 nd 43, respectively), displyed ovious shoulders t dp 18 21. The reltively high rtio of the intensities of pek 1 to the shoulder suggested defective crystlline structure, which could contriute to the very low geltiniztion temperture. Potto strch hd long rnches with n verge rnch chin length of dp 29.4, ut this strch contined the highest concentrtion of phosphte monoester derivtives. The phosphte monoester derivtives, in ddition to hving B- polymorph, contriuted to the low geltiniztion temperture. With the sme chin length, strch of the B-polymorph disply lower geltiniztion temperture thn tht of the A-polymorph (Whittn et l 1990). This chrcteristic could e ttriuted to the presence of 36 molecules of wter in unit cell of the B-polymorph (Srko nd Wu 1978, Imerty et l 1991). Strch of the A-polymorph is more densely pcked with 8 wter molecules in unit cell (Srko nd Wu 1978). Strches tht displyed high geltiniztion tempertures, such s Chinese tro, green nn, cttil millet, norml rice, e wxy, nd high-mylose mize V nd VII, in generl, displyed no shoulder or shoulder of very high reltive intensity (e.g., e wxy nd high-mylose mize strches). Geltiniztion temperture rnges of strches vried from 6.6 C (rley) to 58.8 C (high-mylose mize VII) (Tle IV). Cution is needed for the study of strch geltiniztion properties ecuse mny growing nd processing conditions cn chnge the geltiniztion temperture of strch. Different methods used for strch isoltion were oserved to ffect the geltiniztion temperture of strch. For exmple, wxy mrnth strch isolted y strong-lkline method (Myers nd Fox 1994) displyed significntly higher geltiniztion temperture (70.4 C) thn the 66.7 C for strch isolted from the sme smple y mild lkline nd enzymtic method (Rdosvljevic et l 1998). Wxy strches re known to disply lrger geltiniztion enthlpy chnges, reflecting higher percentge crystllinity of mylopectin. Strches with longer rnch chin length, such s high-mylose mize V nd VII, e wxy mize, nd potto, lso displyed lrger enthlpy chnges, indicting lrger mounts of energy were needed to geltinize crystllites of longer chin lengths. Strch Retrogrdtion DSC dt of strch retrogrdtion re summrized in Tle V. Onset therml trnsition tempertures of dissociting retrogrded strches fter storge t 4 C for seven dys rnged etween 37.3 nd 46.6 C nd were lower thn the onset geltiniztion temperture of ntive strches (56.9 71.5 C). The percentge retrogrdtion (%R), clculted y H retrogrded strch / H geltiniztion, rnged from 4.3% for sweet rice strch to 80.8% for high-mylose mize V. Strches tht displyed the lowest retrogrdtion rtes were wxy rice, sweet rice, nd wxy mrnth; however, not ll wxy strches displyed low retrogrdtion rtes. Wxy mize strch displyed retrogrdtion rte similr to tht of norml mize strch ( 47%); du wxy mize nd e wxy mize strches oth displyed mong the highest retrogrdtion rtes (71.2 nd 61.6%, respectively). Strch of e wxy mize with very long rnch chin length (verge dp 29.5) nd the longest second pek chin length (dp 53) ws expected to hve high retrogrdtion rte. Interestingly, du wxy mize strch hd reltively short verge chin length (dp 23.1) compred with wxy mize strch (dp 23.5) ut hd very long second pek chin length (dp 51) nd the lowest proportion (1.21%) of short chins of dp 6 9. 31 P-NMR studies hve shown tht du wxy mize strch contins phospholipids, which is rre mong wxy strches (Lim et l 1994). Retrogrdtion rtes of strches were inversely correlted with the proportion of short chins of dp 6 9 s proposed y Shi nd Sei (1992), ut no correltion ws found with the molr proportion of unit chins with dp 14 24. Results suggested tht mylopectin molecules with rnched structures cn e eqully effective for crystlline structure formtion, depending on the structure of rnch chins. Fig. 2. Amylopectin rnch chin-length distriutions of B-type strches determined y using high-performnce nion-exchnge chromtogrphy system equipped with n enzyme column rector nd pulsed mperometric detector (HPAEC-ENZ-PAD). A Cropc PA100 column nd n immoilized myloglucosidse column were used for the nlysis. Vol. 76, No. 5, 1999 633
Results of this study showed tht cerel strches, in generl, retrogrded more rpidly thn the tuer nd root strches. This could e ttriuted to the presence of phosphte monoesters in the tuer nd root strches (Lim et l 1994), which retrd retrogrdtion. In contrst, the presence of lipids nd phospholipids in cerel strches nd some wxy cerel strches (e.g., du wxy mize strch) (Lim et l 1994), which restrict the strch grnule swelling nd dispersion, expedite retrogrdtion. Fig. 3. Amylopectin rnch chin-length distriutions of C-type strches determined y using high-performnce nion-exchnge chromtogrphy system equipped with n enzyme column rector nd pulsed mperometric detector (HPAEC-ENZ-PAD). A Cropc PA100 column nd n immoilized myloglucosidse column were used for the nlysis. Psting Properties Psting viscosity profiles of strches nlyzed y using RVA re shown in Fig. 4, nd results re summrized in Tle VI. Psting properties of strch re ffected y mylose nd lipid contents nd y rnch chin-length distriution of mylopectin. Amylopectin contriutes to swelling of strch grnules nd psting, wheres mylose nd lipids inhiit the swelling (Tester nd Morrison 1990). Furthermore, the mylopectin chin-length nd mylose moleculr size produce synergistic effects on the viscosity of strch pstes (Jne nd Chen 1992). Thus, the effects of the structurl fetures on the psting properties of strches re rther complex. The RVA results showed tht the psting tempertures of ll strches were higher thn the onset geltiniztion tempertures (T o ) determined y DSC. The differences rnged etween 3 C of wxy mrnth nd tpioc strches nd >30 C of whet nd rley strches. In generl, wxy cerel strches (except du wxy mize nd e wxy mize) hd lower psting tempertures, higher pek viscosity, nd lower set-ck viscosity thn the norml strch counterprts (Fig. 4 nd nd Tle VI). Wxy strches consisted minly of mylopectin nd, thus, the swelling of grnules ws not restricted y mylose-lipid complex. With no mylose present in the wxy strches exmined in this study, set-ck viscosity, which reflects gel network formtion involving mylose, ws generlly low. Strches of du wxy nd e wxy mize (Fig. 4), however, hd different psting profiles from the forementioned wxy strches. Both strches hd reltively high psting tempertures (75.7 nd 83.2 C, respectively), lower pek viscosity (109 nd 162 RVU, respectively) nd smller rekdown in viscosity during the holding period t 95 C. It is plusile tht the very long rnch-chins of mylopectin mimic mylose to form helicl complexes with lipids nd intertwine with other rnch chins to hold the integrity of strch grnules during heting nd shering. The mylose-lipid complexes in norml strches, such s norml mize (Fig. 4), norml rice (Fig. 4), whet, nd rley (Fig. 4c), cused n increse in psting temperture nd incresed resistnce to sher-thinning of strch pstes. This effect hs een more pronounced in whet nd rley strches ecuse oth hve high concentrtions of phospholipids (0.053% phosphorus in the whet [Lim et l 1994] nd 0.052% in the rley [Song nd Jne, in press]). 634 CEREAL CHEMISTRY TABLE IV Therml Properties of Strch Geltiniztion Determined y Differentil Scnning Clorimetry Type T o ( C) T p ( C) T c ( C) Rnge ( C) H (J/g) A-type strch Norml mize 64.1 ± 0.2 69.4 ± 0.1 74.9 ± 0.6 10.8 12.3 ± 0.0 Wxy mize 64.2 ± 0.2 69.2 ± 0.0 74.6 ± 0.4 10.4 15.4 ± 0.0 du Wxy mize 66.1 ± 0.5 74.2 ± 0.4 80.5 ± 0.2 14.4 15.6 ± 0.2 Norml rice 70.3 ± 0.2 76.2 ± 0.0 80.2 ± 0.0 9.9 13.2 ± 0.6 Wxy rice 56.9 ± 0.3 63.2 ± 0.3 70.3 ± 0.7 13.4 15.4 ± 0.2 Sweet rice 58.6 ± 0.2 64.7 ± 0.0 71.4 ± 0.5 12.8 13.4 ± 0.6 Whet 57.1 ± 0.3 61.6 ± 0.2 66.2 ± 0.3 9.1 10.7 ± 0.2 Brley 56.3 ± 0.0 59.5 ± 0.0 62.9 ± 0.1 6.6 10.0 ± 0.3 Wxy mrnth 66.7 ± 0.2 70.2 ± 0.2 75.2 ± 0.4 8.5 16.3 ± 0.2 Cttil millet 67.1 ± 0.0 71.7 ± 0.0 75.6 ± 0.0 8.5 14.4 ± 0.3 Mung en 60.0 ± 0.4 65.3 ± 0.4 71.5 ± 0.4 11.5 11.4 ± 0.5 Chinese tro 67.3 ± 0.1 72.9 ± 0.1 79.8 ± 0.2 12.5 15.0 ± 0.5 Tpioc 64.3 ± 0.1 68.3 ± 0.2 74.4 ± 0.1 10.1 14.7 ± 0.7 B-type strch e Wxy mize 71.5 ± 0.2 81.0 ± 1.7 97.2 ± 0.8 25.7 22.0 ± 0.3 Amylomize V 71.0 ± 0.4 81.3 ± 0.4 112.6 ± 1.2 41.6 19.5 ± 1.5 Amylomize VII 70.6 ± 0.3 nd 129.4 ± 2.0 58.8 16.2 ± 0.8 Potto 58.2 ± 0.1 62.6 ± 0.1 67.7 ± 0.1 9.5 15.8 ± 1.2 Green lef cnn 59.3 ± 0.3 65.4 ± 0.4 80.3 ± 0.3 21.0 15.5 ± 0.4 C-type strch Lotus root 60.6 ± 0.0 66.2 ± 0.0 71.1 ± 0.2 10.5 13.5 ± 0.1 Green nn 68.6 ± 0.2 72.0 ± 0.2 76.1 ± 0.4 7.5 17.2 ± 0.1 Wter chestnut 58.7 ± 0.5 70.1 ± 0.1 82.8 ± 0.2 24.1 13.6 ± 0.5 Onset temperture (T o ), pek temperture (T p ), completion temperture (T c ), nd enthlpy chnge ( H) of strch geltiniztion. Rnge of geltiniztion is T c T o. Vlues re verges of t lest three replictes of ech smple. Not detectle.
Therefore, whet nd rley strches hd the highest psting tempertures nd very low pek viscosity (Tle VI). Becuse of very high geltiniztion tempertures (>100 C), high-mylose mize V nd VII strches did not completely geltinize under the RVA cooking conditions. Both strches hve very high mylose contents (Tle I) tht did not contriute to swelling, nd thus displyed very low viscosity s shown in Fig. 4. Cttil millet strch hd the lowest solute mylose content (15.3 %) mong norml cerel strches. This feture might explin its high pek viscosity (201 RVU) nd low resistnce to sher-thinning (hot pste viscosity, 80 RVU). In ddition to the effects of mylopectin, mylose, nd lipids, the study lso showed tht different methods used for strch isoltion ffected the psting properties. Exmples were wxy mrnth I TABLE V Therml Properties of Strch Retrogrdtion Determined y Differentil Scnning Clorimetry Type T o ( C) T p ( C) T c ( C) H (J/g) %R A-type strch Norml mize 39.0 ± 0.3 50.1 ± 0.6 59.4 ± 0.1 5.8 ± 0.5 47.6 Wxy mize 40.2 ± 0.5 51.3 ± 0.0 60.2 ± 0.2 7.3 ± 0.2 47.0 du Wxy mize 37.3 ± 0.1 49.4 ± 0.4 60.4 ± 0.2 11.1 ± 0.2 71.2 Norml rice 40.3 ± 0.4 51.0 ± 0.3 60.4 ± 0.2 5.3 ± 0.2 40.5 Wxy rice 43.2 ± 2.1 50.6 ± 0.1 55.2 ± 1.1 0.8 ± 0.5 5.0 Sweet rice 39.8 ± 0.7 50.5 ± 0.0 54.9 ± 0.5 0.6 ± 0.0 4.3 Whet 38.6 ± 0.3 47.7 ± 0.9 55.7 ± 0.2 3.6 ± 0.1 33.7 Brley 39.9 ± 0.8 48.7 ± 0.1 56.5 ± 0.2 3.3 ± 0.5 32.5 Wxy mrnth 42.8 ± 0.4 50.7 ± 0.0 55.0 ± 0.4 0.8 ± 0.2 5.2 Cttil millet 40.5 ± 0.7 50.5 ± 0.0 60.0 ± 0.5 7.8 ± 0.0 53.8 Mung en 39.0 ± 0.7 50.3 ± 0.5 62.3 ± 1.2 6.7 ± 0.8 58.9 Chinese tro 40.8 ± 0.7 51.2 ± 0.2 57.9 ± 0.2 4.8 ± 0.2 32.0 Tpioc 42.1 ± 0.4 51.2 ± 0.5 58.8 ± 0.1 3.7 ± 0.0 25.3 B-type strch e Wxy mize 40.1 ± 1.1 63.0 ± 1.9 87.3 ± 0.4 13.6 ± 0.5 61.6 Amylomize V 44.1 ± 0.8 nd 120.5 ± 2.1 15.8 ± 1.2 80.8 Amylomize VII 46.6 ± 0.5 nd 115.4 ± 1.8 9.9 ± 0.5 61.0 Potto 42.5 ± 0.4 55.7 ± 0.6 66.9 ± 1.0 7.5 ± 0.3 43.4 Green lef cnn 41.3 ± 0.3 57.6 ± 0.1 74.0 ± 1.6 7.0 ± 1.2 45.2 C-type strch Lotus root 41.0.± 0.3 51.5 ± 0.0 62.0 ± 0.0 5.8 ± 0.1 43.2 Green nn 40.7 ± 0.2 52.5 ± 0.0 64.4 ± 0.1 8.2 ± 0.1 47.7 Wter chestnut 40.1 ± 0.2 51.3 ± 0.0 63.3 ± 0.2 6.5 ± 0.3 47.9 Onset temperture (T o ), pek temperture (T p ), completion temperture (T c ), nd enthlpy chnge ( H) of dissociting retrogrded strch. %R is precentge of retrogrdtion. Vlues re verges of t lest three replictes of ech smple. Not detectle. TABLE VI Psting Properties of Strches Mesured y Rpid ViscoAnlyser Psting Temperture Viscosity (RVU) Source ( C) Pek Hot Pste Finl Set-Bck Norml mize 82.0 152 95 169 74 Wxy mize 69.5 205 84 100 16 du Wxy mize 75.7 109 77 99 22 e Wxy mize 83.2 162 150 190 40 Wxy rice 64.1 205 84 100 16 Sweet rice 64.6 219 100 128 28 Norml rice 79.9 113 96 160 64 Whet 88.6 104 75 154 79 Brley I c 87.8 58 15 54 39 Brley II c 91.2 88 58 116 58 Wxy mrnth I d 75.0 60 29 35 6 Wxy mrnth II d 70.2 125 75 86 11 Cttil millet 74.2 201 80 208 128 Chinese tro 73.1 171 88 161 73 Tpioc 67.6 173 61 107 46 Lotus root 67.4 307 84 138 54 Potto 63.5 702 165 231 66 Mung en 73.8 186 161 363 202 Green nn 74.0 250 194 272 78 Wter chestnut 74.3 61 16 27 11 Pstes e Norml mize 90.5 28 18 25 17 Potto 64.4 234 94 114 30 Green lef cnn 71.1 183 181 294 113 Mixtures consisted of 8% (w/w, ds) strch in wter unless otherwise stted. Mesured in Rpid ViscoAnlyser units. c Brley I nd rley II were commercil lrge-grnule strch nd lortory-isolted strch, respectively. d Wxy mrnth I nd II were isolted y the lkline method (Myers nd Fox 1994) nd the diluted lkline nd protese method (Rdosvljevic et l 1998), respectively. e Pstes consisted of 5% strch (w/w, ds). Vol. 76, No. 5, 1999 635
nd wxy mrnth II, isolted y strong lkline method (Myers nd Fox 1994) nd y low lkline-protese method (Rdosvljevic et l 1998), respectively. Psting profiles of the two strches (Fig. 4c) showed tht the mrnth I strch, isolted y the strong lkline method, displyed sustntilly lower viscosity nd higher psting temperture, indicting dmged strch. Among tuer strches, potto strch hd n extrordinrily high pek viscosity (702 RVU, Fig. 4d) s result of the high phosphte monoester content (0.089%, dry sis) (Lim et l 1994; McPherson Fig. 4. Psting profiles of strches (w/w, ds) mesured y rpid visconlysis. A, Mize mutnts t 8% strch. B, Rice mutnts t 8% strch. C, Other cerel strches t 8% strch. Wxy mrnth I nd II strches were prepred y n lkline method (Myers nd Fox 1994) nd the diluted lkline nd protese method (Rdosvljevic et l 1998), respectively. Brley I nd II strches were commercil lrge grnule strch nd lortory-isolted glcier rley strch, respectively. D, Root nd tuer strches t 8% strch. E, Potto, green lef cnn, nd norml mize t 5% strch. 636 CEREAL CHEMISTRY
nd Jne 1999) nd long rnch chins. Compred with norml cerel strches, tuer nd root strches hd lower psting tempertures, lower resistnce to sher-thinning, nd lower set-ck viscosities. These properties were ttriuted to the sence of lipids nd phospholipids in the tuer nd root strches (Lim et l 1994). Among the tuer nd root strches shown in Fig. 4d, Chinese tro strch hd the highest psting temperture nd the lest sher thinning. These fetures my relte to its smll strch grnules (1 4 µm, dimeter) (Jne et l 1994). Green lef cnn strch exhiited very high pek viscosity (183 RVU t 5% concentrtion) compred with norml mize strch (28 RVU) nd potto strch (234 RVU) t the sme concentrtion (Fig. 4e). The strch pste of green lef cnn did not show rekdown in viscosity during the holding period t 95 C nd hd much higher set-ck viscosity (113 RVU) thn potto strch (30 RVU) nd norml mize strch (16 RVU). This unique psting profile of green lef cnn strch could e ttriuted to the presence of high mylose content nd high proportions of very long mylopectin rnch chins. Mung en strch, consisting of high solute-mylose content (30.7%) lso displyed very high set-ck viscosity (202 RVU) (Tle VI). CONCLUSIONS The studies showed tht mylopectin rnch chin lengths nd distriutions determined strch geltiniztion temperture, enthlpy chnge, nd psting properties. Strch geltiniztion temperture incresed with incresing rnch chin-length. Those strches displying sustntil shoulder t dp 18 21 in the rnch chinlength distriution, in generl, hd lower geltiniztion tempertures. Incresing mylose contents, long with lipids nd phospholipids, significntly incresed strch psting temperture, decresed pek viscosity nd sher thinning, nd incresed set-ck viscosity. The very long rnch-chins of mylopectin (dp > 73), like mylose, could ind iodine nd inflte the iodine ffinity. The very long rnch-chins, like mylose, lso ffected strch-psting properties. Strches isolted y different methods displyed different physicl properties. ACKNOWLEDGMENTS We thnk the Iow Corn Promotion Bord, Pioneer Hi-Bred Interntionl, Inc. nd Du Pont Agriculturl Products, for finncil support; Y. Song, S. H. Yoo, nd L. F. Hnson for providing lortory isolted rley strch nd mrnth strch, respectively; nd N. Morin for editoril service. LITERATURE CITED Asok, M., Okuno, K., nd Fuw, H. 1985. Effect of environmentl temperture t the milky stge on mylose content nd fine structure of mylopectin of wxy nd nonwxy endosperm strches of rice. Agric. Biol. Chem. 49:373-379. Cmeron, R. E., nd Donld, A. M. 1992. A smll-ngle X-ry scttering study of the nneling nd geltiniztion of strch. Polymer 33:2628-2635. Gidley, M. J., nd Bulpin, P. V. 1987. Crystlliztion of mltooligoscchrides s models of the crystlline forms of strch: Minimum chin-length requirement for the formtion of doule helices. Crohydr. 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Lnsky, S., Kooi, M., nd Schoch, T. J. 1949. Properties of the frctions nd liner sufrctions from vrious strches. J. Am. Chem. Soc. 71:4066-4075. Lim, S. T., Ksemsuwn, T., nd Jne, J. 1994. Chrcteriztion of phosphorus in strches using 31 P-NMR spectroscopy. Cerel Chem. 7:488-493. Lu, S., Chen, L.-N., nd Lii, C.-Y. 1997. Reltions etween the fine structure, physicochemicl properties, nd retrogrdtion of mylopectins from Tiwn rice vrieties. Cerel Chem. 74:34-39. McPherson, A. E., nd Jne, J. 1999. Physiclchemicl properties of selected root nd tuer strches. Crohydr. Polym. 40:57-70. Miles, M. J., Morris, V. J., Orford, P. D., nd Ring, S. G. 1985. The roles of mylose nd mylopectin in the geltion nd retrogrdtion of strch. Crohydr. Res. 135:271-281. Morrison W. R., nd Lignelet, B. 1983. An improved colorimetric procedure for determining pprent nd totl mylose in cerel nd other strches. J. Cerel Sci. 1:9-20. Myers, D. J., nd Fox, S. R. 1994. Alkli wet-milling chrcteristics of perled nd unperled mrnth seed. Cerel Chem. 71:96-99. Rdosvljevic, M., Jne, J., nd Johnson, L. A. 1998. Isoltion of mrnth strch y diluted lkline-protese tretment. Cerel Chem. 75:212-216. Srko, A., nd Wu, H.-C. H. 1978. The crystl structures of A-, B-, nd C-polymorphs of mylose nd strch. Strch/Sterke 30:73-78. Shi, Y. C., nd Sei, P. A. 1992. The structure of four wxy strches relted to geltiniztion nd retrogrdtion. Crohydr. Res. 227:131-145. Shi, Y. C., nd Sei, P. A. 1995. Fine structure of mize strches from four wx-contining genotypes of the W64A inred line in reltion to geltiniztion nd retrogrdtion. Crohydr. Polym. 26:141-147. Song, Y., nd Jne, J. In press. Chrcteriztion of rley strches from wxy, norml nd high mylose vrieties. Crohydr. Polym. Tked, Y., nd Hizukuri, S. 1987. Structures of rice mylopectins with low nd high ffinities for iodine. Crohydr. Res. 168:79-88. Tester, R. F., nd Morrison, W. R. 1990. Swelling nd geltiniztion of cerel strches. I. Effects of mylopectin, mylose, nd lipid. Cerel Chem. 67:551-557. Wng, Y. J., White, P. J., nd Pollk, L. 1993. Physicochemicl properties of strches from mutnt genotypes of the Oh43 inred line. Cerel Chem. 70:199-203. Whittm, M. A., Noel, T. R., nd Ring, S. G. 1990. Melting ehviour of A- nd B-type crystlline strch. Int. J. Biol. Mcromol. 12:359-362. Wong, K. S., nd Jne, J. 1997. Quntittive nlysis of dernched mylopectin y HPAEC-PAD with post-column enzyme rector. J. Liq. Chrom. 20:297-310. Yun, R. C., Thompson, D. B., nd Boyer, C.D. 1993. Fine structure of mylopectin in reltion to geltiniztion nd retrogrdtion ehvior of mize strches from three wx-contining genotypes in two inred lines. Cerel Chem. 70:81-89. [Received Ferury 10, 1999. Accepted June 10, 1999.] Vol. 76, No. 5, 1999 637