preparation of nonfragmented, completely amorphous, pregelatinized maize starches and determination...

Irma AmeliaJames N. BeMiller

Whistler Center for CarbohydrateResearch,Purdue University,West Lafayette, IN, USA

Preparation of Nonfragmented, CompletelyAmorphous, Pregelatinized Maize Starches andDetermination of the Effects of Fragmentation onthe Adhesiveness of Their Pastes

The objective of this research was to prepare pregelatinized maize starches that wereboth completely amorphous (no crystalline structure) and unfragmented, and to applythe same method to partially depolymerized maize starches to determine the effect offragmentation on the degree of stickiness (tackiness, adhesiveness). Amorphouspregelatinized normal maize starch could be prepared by precipitation of a paste withacetone. Precipitation with a polar organic solvent was not applicable for the prepara-tion of amorphous, pregelatinized waxy maize starch, because a sticky, cohesive andadhesive mass was produced. Therefore, freeze drying was used to prepare amor-phous pregelatinized waxy maize starch. Limited acid-catalyzed hydrolysis of normalmaize starch before pregelatinization increased stickiness up to a maximum, afterwhich the degree of stickiness decreased. In all aspects of this research, it was clearthat the behavior of waxy maize starch was rather different than that of normal maizestarch.

Keywords: Amorphous maize starch; Starch paste adhesiveness; Starch pastestickiness

696 Starch/Stärke 61 (2009) 696–701

1 Introduction

Pregelatinized starches are precooked and dried star-ches, whose methods of preparation involve more thangelatinization of the granules. Two basic processes areused to prepare conventional pregelatinized starches,i.e., starch products in which few, if any, granules, in eitherbirefringent or non-birefringent forms, are present. In one,a starch-water slurry is applied to a drum heated withsuperheated steam. The starch is cooked instantaneouslyto a paste that dries rapidly (before the drum makes a fullturn). The dried film is scraped off and ground to a powder[1-4]. Another process uses some form of extrusion [2, 3,5]. Starch and water are introduced into one end of anextruder. Dry, puffed material exits the other end. Thismaterial is also ground to a powder. This paper is relatedto these two conventional pregelatinized starches.

Starches are somewhat depolymerized as a result of theheat and/or shear they are subjected to in the process.For example, the molecular weights of wheat starchamylose and amylopectin were reported to decrease byfactors of 1.1 and 2.6, respectively, during a drum cooking

and drying process and by factors of 1.5 and 15, respec-tively, during an extrusion process [2, 3].

Isolation and characterization of a soluble, branchedstarch fraction from maize masa indicated that molecularfragments of amylopectin produced by milling are relatedto stickiness in food products [6]. One of us (J.N.B.) is aparticipant in a project in which a similar phenomenon,i.e., fragmentation of amylopectin molecules, has beenfound in commercial pregelatinized starches, presumablyas a result of milling of flakes from the hot drum or of heatand shear the moist starch is subjected to in an extruderand/or the milling of the puffed product. The paper byColonna et al. [2] states that the studied pregelatinizedwheat starch preparations, which had been cooked anddried on a drum or that had been prepared in a extruder,were dried in a vacuum oven at 457C for seven days, thenground and sieved to give a particle size of .50 mm.Grinding and sieving is not mentioned in the article byDoublier et al. [3]. In the studies of Colonna et al. [2] andby Doublier et al. [3], it is not known to what extent starchmolecule depolymerization may have occurred during thepregelatinization process and/or during milling. This studyis designed to provide some insight into this question.

That starch is labile to thermal degradation was found byHan et al. [7]. Han and Lim [8] also found that depolymer-ization occurs during excessive vortexing. Barth and

Correspondence: James N. BeMiller, Purdue University – FoodScience, 745 Agriculture Mall Dr, West Lafayette, Indiana 47907-2009, USA. E-mail: [email protected].

© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.starch-journal.com

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DOI 10.1002/star.200900160

Starch/Stärke 61 (2009) 696–701 Nonfragmented, Completely Amorphous, Pregelatinized Maize Starches 697

Carlin [9] found that passage of a starch solution througha high-pressure, size-exclusion chromatography columncan effect depolymerization, which is why a medium-pressure column was used in this work and that of Miklusand Hamaker [6] and Han et al. [7].

We wanted to determine how molecular fragmentation isrelated to stickiness (tackiness, adhesiveness) of pastesand gels made from pregelatinized maize starches. How-ever, there was evidence (from the other project) that thecontent of ungelatinized granules and granule fragmentsvaries with the parent starch and the process used (hotdrum vs. some type of extrusion). The goal, therefore, wasto fragment normal and waxy maize starch in a controlledway and then to produce completely amorphous prod-ucts, i.e., starch devoid of granules, granule fragments,and crystallinity, from both the acid-treated and intactstarches. Because Miklus and Hamaker [6] had con-cluded that the component causing stickiness was com-posed of amylopectin fragments, an all-amylopectinstarch (waxy maize starch) was used along with normalmaize starch.

2 Materials and Methods

2.1 Preparation of amorphous normal maizestarch

All water used was deionized, then glass-distilled. Normalmaize starch (Pure Food Powder, Tate & Lyle NorthAmerica, Decatur, IL, USA) was pasted by heating a 5%slurry in water at 1007C for 10 min with continuous mod-erate stirring with a mechanical stirrer. The paste was thenallowed to cool to ,707C while stirring was continued.The rate of stirring was increased slightly, and threevolumes of ethanol were added slowly. The precipitatewas collected by centrifugation, dehydrated by washingwith 95% 2-propanol, followed by 100% 2-propanol, andair dried. This preparation was redissolved in water (at aconcentration of 5%) and heated at the boiling tempera-ture for 10 min with moderate stirring. The solution wasallowed to cool to ,707C, then three volumes of acetonewere added gradually while stirring was continued. Theprecipitate was collected by centrifugation, dehydratedby washing three times with acetone, and air dried. Thecake was gently ground using a mortar and pestle andpassed through a 44-mm sieve.

2.2 Preparation of amorphous waxy maizestarch

Waxy maize starch (Amioca TF, National Starch andChemical Co., Bridgewater, NJ, USA) was pasted by

heating a 5% slurry in water at 1007C for 10 min withcontinuous moderate stirring with a mechanical stirrer.The hot paste was then frozen rapidly as a shell in Lab-conco freeze-dryer flasks using a mixture of acetone andsolid carbon dioxide (-787C) and freeze dried (LabconcoCorp., Kansas City, MO, USA).

2.3 RVA analysis

Pasting and paste properties of the amorphous starches,before and after depolymerization, were determinedusing a Rapid Visco Analyser (Model RVA-4C, NewportScientific Pty. Ltd., Warriewood, Australia). Slurries (25 g,8%, w/w) were stirred for 1 h at room temperature, thenpoured into aluminum canisters and stirred manuallyusing the plastic paddles. Pasting characteristics weredetermined with the Rapid Visco Analyser using standardprofile 1. A 13-min analysis was used: equilibration to507C for 60 s, heating to 957C in 222 s, holding at 957C for150 s, cooling to 507C in 228 s, holding at 507C for 120 s.

2.4 Depolymerization/fragmentation of thestarches

Slurries (40%) of granular normal and waxy maize star-ches in 0.5 M aqueous HCl were heated at 507C for var-ious times. The slurries were then neutralized to pH 6.0-6.5 with 1.2 M aqueous NaOH. The acid-modified star-ches were isolated by centrifugation, washed three timeswith water, and air dried before being converted to amor-phous products as described in Sections 2.1 and 2.2.

2.5 Size-exclusion chromatographic analysis

Amorphous starch (100 mg) from control and acid-thinned preparations was mixed with 1.0 mL of water in acentrifuge tube. Then, 9.0 mL of dimethyl sulfoxide wasadded, and the tube was placed in a boiling water bath for1 h with occasional vortexing. The solution was cooled toroom temperature. Then, three volumes of ethanol wereadded gradually and with stirring (magnetic stir bar). Theprecipitate was collected by centrifugation (28006g, 25min). The supernatant was discarded; the precipitate wasdissolved in water and reprecipitated with three volumesof ethanol for three additional times. The precipitate wasfinally dehydrated with 100% ethanol and air dried. Waterwas added to the dry sample so that a concentration of,5 mg/mL was obtained. The centrifuge tube was thenplaced in a boiling water bath for 30 min and vortexedfrequently. This solution was loaded onto a calibratedcolumn of Sephacryl S-500 HR (Amersham Biosciences,Piscataway, NJ, USA) through a 200-mL loop injector


698 I. Amelia and J. N. BeMiller Starch/Stärke 61 (2009) 696–701

(Rheodyne 7125, Cotati, CA, USA), passing through a 5.0-mm nylon filter during injection. The column was con-nected to refractive index (Wyatt Optilab 903) and MALLSlaser photometer (DAWN D SP-F, wavelength 488 nm witha K-5 flow cell [Wyatt Technology, Santa Barbara, CA,USA]) detectors. A degassed, filtered (0.1-mm Milliporefilter) aqueous mobile phase of 0.02% sodium azide at aflow rate of 1.3 mL/min was used for elution. The data wasprocessed using ASTRA for windows software (version4.90. U8, Wyatt Technology Corp., Santa Barbara, CAUSA) and the Berry extrapolation method.

2.6 Texture profile analysis

Texture profile analysis was done using a TA-XT PlusTexture Analyzer (TA Instrument Co., Cincinnati, OH,USA). Amorphous and control starches were addedgradually to continuously stirred water (magnetic stir bar)in a glass beaker. A concentration of 12% was used fornormal maize starch and its products and a concentrationof 14% for waxy maize starch and its products. The mix-ture was stirred until a homogeneous system was formed.The beaker was covered with Parafilm (Structure Probe,Inc., West Chester, PA, USA) and let stand for 120 min.The experimental conditions were: probe P/O. 245, 0.25in, spherical, stainless steel; test speed, 3 mm/s; post-test speed, 3 mm/s; target mode, distance; time, 3 s (resttime between cycles); trigger type, Lutton; tare mode, on;advance options, on.

2.7 X-ray diffraction

The starch samples (,500 mg) were back-pressed intoaluminum holders and mounted in a Philips PW3710 dif-fractometer interfaced to a personal computer equippedwith Automated Powder Diffraction (APD) software (Phi-lips/PANalytical B.V., Almelo, The Netherlands). Ni-filteredCu Ka radiation (l = 0.1518 nm) was used, and the tubewas operated at 40 kV and 25 mA. The intensity data werecollected at room temperature in 0.027 intervals over the2y range 77- 367. The time spent at each step was 5 s. Thepatterns were smoothed for further analysis by the PC-APD (version 3.6) software.

3 Results and Discussion

The basic idea was to precipitate the starch from a freshlyprepared hot paste using a water-soluble organic solventbefore the starch molecules had a chance to retrograde/crystallize. Normal maize starch (NMS) was used first. X-ray diffraction analysis revealed that precipitation byethanol and 2-propanol resulted in formation of some

starch-organic solvent complexes and some crystallinity.Precipitation with acetone produced a completely amor-phous starch. However, the starch was first precipitatedwith ethanol because absence of this step resulted in asticky and cohesive mass. When the ethanol precipitatewas washed with 2-propanol, dried, redissolved in water,and precipitated with acetone, a fluffy powder thatexhibited no peaks indicative of crystallinity in an X-raydiffractogram was obtained (Fig. 1a).

The procedure used with normal maize starch was notsuitable for waxy maize starch (WMS). All attempts toapply it to WMS resulted in a sticky mass that could bedehydrated, but the resulting product was a hard massthat had to be ground, something we wanted to avoid.However, it was found that rapid freezing of a hot pasteand freeze-drying gave amorphous amylopectin (AP)(Fig. 1b).

It was then found that amorphous starch could not beeasily milled, except possibly in a ball mill. So to givecontrolled depolymerization, mild acid-catalyzed hy-drolysis was used to degrade the starch polymers to aslight extent. RVA analysis data for the control and acid-thinned starches are given in Tab. 1.

SEC results revealed that the molecular weight of AP inhydrolyzed normal maize starch was more extensivelyreduced than that in hydrolyzed waxy maize starch. Theratio between the area under the first peak (largermolecules [amylopectin] that eluted at 36-48 mL, re-ferred to as Area 1) and the area under the broad sec-ond peak (smaller molecules [amylose and amylopectinfragments] that eluted at 48-90 mL) was determined(Tab. 2). In both cases, it is clear that AP moleculeswere converted into smaller molecules whose elution(and size) overlapped that of AM molecules as a func-tion of hydrolysis time.

The stickiness/tackiness/adhesiveness of the variousproducts was determined by texture profile analysis(TPA). Due to differences in paste/gel hardness, thepeak forces of both controls (waxy and normal cornstarch) were significantly greater than was that of thehydrolyzed samples when an equal degree of compres-sion was applied. The more hydrolyzed was the starchthe lower was the peak force. The negative area in theTPA graph, which was a measure of adhesiveness/tackiness/stickiness, correlated to the level of peakforce. In order to overcome the differences in paste/gelhardness, the data was normalized by adjusting thedegree of compression so that approximately equalpeak force was achieved, and the respective negativearea was measured (Tab. 3).



Fig. 1. X-ray diffraction pattern of (a) amor-phous normal maize starch and (b) amor-phous waxy maize starch.

For NMS, hydrolysis increased paste adhesiveness up toa maximum that was about 80-fold greater than the valuefor intact amorphous NMS, after which the degree ofadhesiveness decreased (Tab. 2), data which seems tosupport the hypothesis that a relatively small degree ofdepolymerization was achieved during preparation ofpregelatinized starch, and that that small degree ofhydrolysis resulted in an increase in adhesiveness. How-ever, the same was not true of WMS. A paste of non-depolymerized amorphous WMS had the greatest degreeof adhesiveness (Tab. 2). Any degree of depolymerizationresulted in lower paste adhesiveness values. BecauseMiklus and Hamaker [6] found that fragmentation of AP inmasa made from normal maize starch was the factorrelated to stickiness, we can only conclude that frag-mentation of AP increases stickiness only when AM ispresent.

Overall, the results indicate that, not only do AP frag-ments produce stickiness in food products [6] when in thepresence of AM or AM fragments, but intact AP moleculesmight also contribute to stickiness.

4 Conclusions

(1) Amorphous normal maize starch can be made byprecipitation from a hot paste of completely gelatinizedstarch using acetone as the precipitant. (2) Amorphouswaxy maize starch can be made by freeze drying apaste of completely gelatinized starch. (3) Paste sticki-ness is related to fragmentation of amylopectin, but onlywhen amylose (or partially degraded amylose) is pres-ent.


700 I. Amelia and J. N. BeMiller Starch/Stärke 61 (2009) 696–701

Tab. 1. RVA characteristics for control and hydrolyzed maize starches.

Sample PVa

[RVU]bHPV[RVU]

CPV[RVU]

Breakdown[RVU]

Setback[RVU]

PT[7C]

Normal maize starchControlc 1173 952 1123 221 171 861 h heating in 0.5 M HCl 557 154 238 403 84 752 h heating in 0.5 M HCl 222 27 55 195 28 773 h heating in 0.5 M HCl 90 3 23 87 20 Errd

Waxy maize starchControlc 2298 1504 1852 794 348 751 h heating in 0.5 M HCl 582 137 161 445 24 733 h heating in 0.5 M HCl 50 7 13 43 6 754 h heating in 0.5 M HCl 24 21 10 25 11 Errd

a PV, peak viscosity; HPV, hot paste viscosity; CPV, cold paste viscosity; PT, pasting temperature;RVU, Rapid Visco Analyser units.

b 1 RVU = 12 mPa?s.c Amorphous starch without an acid treatment.d No value recorded.

Tab. 2. Size-exclusion chromatography data.

Sample Area I/Area IIa

Normal maize starchControlb 1.71 h heating in 0.5 M HCl 1.22 h heating in 0.5 M HCl 0.43 h heating in 0.5 M HCl 0.2

Waxy maize starchControlb 5.41 h heating in 0.5 M HCl 6.32 h heating in 0.5 M HCl 4.73 h heating in 0.5 M HCl 0.9

a See text.b Amorphous starch without an acid treatment.

Tab. 3. Texture profile analysis data.

Sample Deg. ofcompression[mm]

Peak forcea

[g]Area ofnegative curvea

[g?s]

Normal maize starchControlb 2.0 10.5 0.531 h heating in 0.5 M HCl 3.2 10.5 16.902 h heating in 0.5 M HCl 5.5 10.9 43.933 h heating in 0.5 M HCl 8.0 10.7 7.28

Waxy maize starchControlb 5.3 9.50 30.131 h heating in 0.5 M HCl 7.5 10.20 12.233 h heating in 0.5 M HCl 8.0 10.20 7.894 h heating in 0.5 M HCl 8.0 9.90 7.41

a Average of 2 determinations.b Amorphous starch without an acid treatment.



Acknowledgement

The authors thank Dr. Srinivas Janaswamy for doing the X-ray diffraction analyses and Xiaohong Sheng for assistingwith the SEC analysis. Author Amelia thanks AgriculturalResearch Programs of the Purdue University College ofAgriculture for an Agricultural Research Fund Scholarshipthat funded this research.

References

[1] E. L. Powell: Production and use of pregelatinized starch, inStarch: Chemistry and Industry. Vol. II (Eds. R. L. Whistler, E.F. Paschall) Academic Press, New York, 1967, pp. 523–536.

[2] P. Colonna, J. L. Doublier, J. P. Melcion, F. Demonredon, C.Mercier: Extrusion cooking and drum drying of wheat-starch.1. Physical and macromolecular modifications. Cereal Chem.1984, 61, 538–543.

[3] J. L. Doublier, P. Colonna, C. Mercier : Extrusion cooking anddrum drying of wheat starch. 2. Rheological characterizationof starch pastes. Cereal Chem. 1986, 63, 240–246.

[4] R. Takahashi, T. Ojima Pregelatinization of wheat starch in adrum drier. Stärke 1969, 21, 318–321.

[5] R. M. L. Alves, M. V. E. Grossman, R. S. S. F. Silva: Gellingproperties of extruded yam (Dioscorea alata) starch. FoodChem. 1999, 67, 123–127.

[6] M. B. Miklus, B. R. Hamaker: Isolation and characterizationof a soluble branched starch fraction from corn masa asso-ciated with adhesiveness. Cereal Chem. 2003, 80, 693–698.

[7] J.-A. Han, J. N. BeMiller, S.-T. Lim: Structural changes ofdebranched corn starch by aqueous heating and stirring.Cereal Chem. 2003, 80, 323–328.

[8] J.-A. Han, S.-T. Lim: Structural changes in corn starchesduring alkaline dissolution by vortexing. Carbohydr. Polym.2004, 55, 193–199.

[9] H. G. Barth, F. J. Carlin: A review of polymer shear degrada-tion in size-exclusion chromatography. J. Liq. Chromatogr1984, 7, 1717–1738.

(Received: November 18, 2008)(Revised: April 7/July 27, 2009)(Accepted: July 27, 2009)


preparation of nonfragmented, completely amorphous, pregelatinized maize starches and determination...

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