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Food Hydrocolloids 23 (2009) 1527–1534

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Food Hydrocolloids

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Form and functionality of starch

Les Copeland*, Jaroslav Blazek, Hayfa Salman, Mary Chiming TangFaculty of Agriculture, Food and Natural Resources, University of Sydney, Sydney, NSW 2006, Australia

a r t i c l e i n f o

Article history:Received 29 August 2008Accepted 21 September 2008

Keywords:StarchGranulesAmyloseAmylopectinFunctional propertiesStarch–lipid complexesEnzymic digestion

* Corresponding author. Tel.: þ61 2 9036 7047; faxE-mail address: l.copeland@usyd.edu.au (L. Copela

0268-005X/$ – see front matter � 2008 Elsevier Ltd.doi:10.1016/j.foodhyd.2008.09.016

a b s t r a c t

Starch is a macro-constituent of many foods and its properties and interactions with other constituents,particularly water and lipids, are of interest to the food industry and for human nutrition. Starch variesgreatly in form and functionality between and within botanical species, which provides starches ofdiverse properties but can also cause problems in processing due to inconsistency of raw materials. Beingable to predict functionality from knowledge of the structure, and explain how starch interacts withother major food constituents remain significant challenges in food science, nutrition, and for the starchindustry generally. This paper describes our current understanding of starch structure that is relevant toits functionality in foods and nutrition. Amylose influences the packing of amylopectin into crystallitesand the organization of the crystalline lamellae within granules, which is important for propertiesrelated to water uptake. Thermal properties and gel formation appear to be influenced by both amylosecontent and amylopectin architecture. While amylose content is likely to have an important bearing onthe functional properties of starch, subtle structural variations in the molecular architecture of amylo-pectin introduces uncertainty into the prediction of functional properties from amylose content alone.Our ability to relate starch granule structure to suitability for a particular food manufacturing process orits nutritional qualities depends not only on knowledge of the genetic and environmental factors thatcontrol starch biosynthesis, and in turn granule morphology, but also on how the material is processed.

� 2008 Elsevier Ltd. All rights reserved.

1. Introduction

Ensuring food security and delivering foods that provide healthbenefits are major global challenges. Increased demand for dietaryanimal protein in Asia, continuing population growth, a worldwideincrease in diet-related illnesses, and competition for arable landand water from alternative crops such as biofuels are pressures thatare driving the need to increase the efficiency and quality of foodproduction. Improving processes and products through ensuringfitness-for-purpose and better use of raw materials will be criticalin achieving these objectives. This requires increasing our under-standing of the relationship between form and functional proper-ties of food constituents.

Most foods are multi-component, multi-phase systems thatcontain complex mixtures of water, polysaccharides, proteins,lipids and numerous minor constituents. Starch is present asa macro-constituent in many foods and its properties and interac-tions with other constituents, particularly water and lipids, are ofinterest to the food industry and for human nutrition. Starchcontributes 50–70% of the energy in the human diet, providinga direct source of glucose, which is an essential substrate in brain

: þ61 2 9351 2945.nd).

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and red blood cells for generating metabolic energy. Indeed, theavailability of a reliable source of starch from agriculture isconsidered to have been an important factor in human develop-ment (Perry et al., 2007), although it now seems that the glycemicresponse to excessive consumption of rapidly digesting starch maybe a factor in some diet-related illnesses. Starch is also an importantindustrial material. Approximately, 60 million tonnes are extractedannually worldwide from various cereal, tuber and root crops, ofwhich roughly 60% is used in foods (for example, bakery products,sauces, soups, confectionery, sugar syrups, ice cream, snack foods,meat products, baby foods, fat replacers, coffee whitener, beer, softdrinks) and 40% in pharmaceuticals and non-edible purposes, suchas fertilisers, seed coatings, paper, cardboard, packing material,adhesives, textiles, fabrics, diapers, bioplastics, building materials,cement, and oil drilling (Burrell, 2003). Starches with a wide rangeof functional properties are needed to ensure fitness-for-purposefor such a diverse range of end uses.

Starch varies greatly in form and functionality between andwithin botanical species, and even from the same plant cultivargrown under different conditions. This variability provides starchesof diverse properties, but it can also cause problems in processingdue to inconsistency of raw materials. As a result, chemicallymodified starches are used extensively to overcome the variabilityof native starches and their lack of versatility over a wide range ofprocessing conditions. However, consumer interest in ‘‘more

L. Copeland et al. / Food Hydrocolloids 23 (2009) 1527–15341528

natural foods’’ is increasing, and hence there is a need for greaterunderstanding of how processing and nutritional performance arerelated to starch morphology.

Starch has been the subject of intensive research over manydecades, resulting in a vast body of published literature onpreparative and analytical methods, molecular structure, physical,chemical and biochemical properties, functionality and uses. Yetbeing able to predict functionality from knowledge of the structure,and explain how starch interacts with other major food constitu-ents remain significant challenges in food science, nutrition, and forthe starch industry generally. It is not intended in this article toreview the extensive literature on starch. Rather, the aim is toprovide the reader with a reasonably concise discussion of ourcurrent knowledge of starch structure that is relevant to its func-tionality in foods and nutrition. The reference articles cited areintended to be illustrative rather than comprehensive. Clearly,many relevant references that could have been included have beenomitted to achieve brevity. For comprehensive treatises on starchstructure, function and applications, the reader is referred to thecompendia edited by Eliasson (2004) and by Yuryev and Tomasik(2007).

2. The form of starch

Starch is the main storage carbohydrate of plants. It is depositedas insoluble, semi-crystalline granules in storage tissues (grains,tubers, roots) and it also occurs to a lesser extent in most vegetativetissues of plants. Starch is made up of two polymers of D-glucose:amylose, an essentially unbranched a[1 / 4] linked glucan, andamylopectin, which has chains of a[1 / 4] linked glucosesarranged in a highly branched structure with a[1 / 6] branchinglinks. The moisture content of native starch granules is usuallyabout 10%. Amylose and amylopectin make up 98–99% of the dryweight of native granules, with the remainder comprising smallamounts of lipids, minerals, and phosphorus in the form of phos-phates esterified to glucose hydroxyls. Starch granules range in size(from 1 to 100 mm diameter) and shape (polygonal, spherical,lenticular), and can vary greatly with regard to content, structureand organization of the amylose and amylopectin molecules, thebranching architecture of amylopectin, and the degree of crystal-linity (Lindeboom, Chang, & Tyler, 2004). Granules may occurindividually or clustered as compound granules, and in wheat,barley, rye and triticale they occur in bimodal size distributions.This diversity in the form of starch granules and their molecularconstituents influences starch functionality.

Although a minor component by weight, lipids can havea significant role in determining the properties of starch. The lipidcontent and composition of starch granules varies among plantspecies. Lipids associated with isolated cereal starch granules occuron the surface as well as inside the granule. Surface lipids aremainly triglycerides, and to a lesser extent free fatty acids, glyco-lipids and phospholipids, which can be extracted with diethylether.Internal lipids of cereal starches are predominantly monoacyl lipidsthat are usually extracted with hot aqueous alcohol. Both surfaceand internal lipids may be present in the free state, linked throughionic or hydrogen bonds to hydroxyl groups, or bound in the formof amylose inclusion complexes (Morrison, 1988; Morrison, 1995;Vasanthan & Hoover, 1992). The lipid content of native starches ishighly correlated with amylose content: the higher the amylosecontent the more lipid is present.

The structure of amylose and amylopectin have been studiedextensively and have been the subject of numerous reviews, assummarized in the following section (for example: Blanshard,1987; Buleon, Colonna, Planchot, & Ball, 1998; Hoover, 2001; Sri-chuwong & Jane, 2007; Tester, Karkalas, & Qi, 2004; Zobel, 1988).Amylose has a molecular weight range of approximately 105–106,

corresponding to a degree of polymerization (DP) of 1000–10,000glucose units. Less than 0.5% of the glucoses in amylose are ina[1 / 6] linkages, resulting in a low degree of branching, anda structure with 3–11 chains of approximately 200–700 glucoseresidues per molecule. Because of the low degree of branching,dissolved amylose has a tendency to form insoluble semi-crystal-line aggregates, depending on the placement of the branches in thestructure.

Amylopectin is a much larger polymer, with molecular weightabout 108 and a DP that may exceed one million. Most starchescontain 60–90% amylopectin, although high-amylose starches, withas little as 30% amylopectin, and waxy starches with essentially100% amylopectin are well known. Amylopectin has about 5% of itsglucoses in a[1 / 6] linkages, giving it a highly branched, tree-likestructure and a complex molecular architecture that can varysubstantially between different starches with regard to placementand length of branches. The amylopectin branches may be classifiedaccording to their pattern of substitution: A-chains are defined asunsubstituted, B-chains are substituted by other chains, and thereis a single C-chain that caries the reducing glucose. Glycogen,a storage polysaccharide of animals, has the same chemicalcomposition as amylopectin but it has a higher degree of branchinggiving it a more compact globular shape. More glucose can bepacked into the open ended, tree-like structure of amylopectin thanin the more closed, globular configuration of glycogen.

The natural variability in amylose and amylopectin molecules isdue to the complexity of starch biosynthesis. Again, the reader isreferred to several excellent in-depth reviews of this topic (Buleonet al., 1998; Morell & Myers, 2005; Smith, Denyer, & Martin, 1997;Tetlow, Morell, & Emes, 2004). The biosynthetic pathway involvesseveral types of enzymes: ADP pyrophosphorylases, which formthe glucosyl donor ADP-glucose from precursor hexose-phos-phates; soluble and granule bound starch synthases, which catalysethe formation of a[1 / 4] linkages; starch branching enzymes,which catalyse the formation of a[1 / 4] branching linkages; andstarch debranching enzymes, which are considered to have a roletrimming newly synthesized amylopectin branches to enable themto pack into crystalline structures. These enzymes occur in multipleforms, and their activities may be subject to temporal and spatialdifferences in expression, environmental influences at both thegenetic and enzyme level, and differences in substrate specificitiesof multiple forms. The end products of the biosynthetic pathwayreflect the genetic diversity among the enzymes involved, andenvironmental influences acting on their expression and activity.The biosynthesis of amylose and the assembly of amylopectin andamylose into the granules are not well understood.

3. Starch granules

The extent of crystallinity of native starch granules ranges fromabout 15% for high-amylose starches to about 45–50% for waxystarches. The granules have a hierarchical structure that can beobserved readily by light and electron microscopy. Multipleconcentric layers of so-called growth rings of increasing diameterextend from the hilum (the centre of growth) towards the surface ofgranules. The growth rings are typically 120–400 nm in thickness,and are considered to represent diurnal fluctuations in the depo-sition of starch in storage tissues (Donald, Kato, Perry, & Waigh,2001; Gallant, Bouchet, & Baldwin, 1997; Ridout, Gunning, Parker,Wilson, & Morris, 2002; Sevenou, Hill, Farhat, & Mitchell, 2002).The concentric growth rings, in turn, contain alternating crystallineand amorphous regions of higher and lower density, respectively.The higher density regions have a lamellar structure of alternatingcrystalline and amorphous layers with a repeat distance of 9–11 nm(Donald, 2004; Donald et al., 2001; Yuryev et al., 2004). Withinthese lamellae, the crystalline layers are considered to be formed

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mainly by amylopectin chains packed into a crystalline lattice,whereas the amorphous layers contain the amylopectin branchingpoints and amylose and amylopectin molecules in a disorderedconformation. Longer amylopectin chains are considered to passfrom the crystalline region into the amorphous region of thelamellae (Jane, 2007; Qi, Tester, Snape, & Ansell, 2003). Amylo-pectin molecules near the surface of granules may have a differentstructure to those closer to the centre of the molecule. The ratio oflong to short branch chains affect the shape of amylopectin, whichinfluences their packing into granules (Jane, 2007). In this way,genetic and environmental factors that affect amylopectin biosyn-thesis can influence its molecular architecture, and in turn, granulemorphology.

Amylose is located in the low density layers of the growth rings,although amylose molecules are also considered to be interspersedbetween amylopectin in the crystalline layers, disrupting the crystalpacking of amylopectin (Hedley, Bogracheva, & Wang, 2002; Jane,2006; Kozlov, Blennow, Krivandin, & Yuryev, 2007; Kozlov, Noda,Bertoft, & Yuryev, 2006; Matveev et al., 1998). In high-amylosestarches, amylose helices may contribute to the crystallinity ofgranules (Buleon et al., 1998; Hoover, 2001; Matveev et al., 2001;Tester et al., 2004). The organization of amylose and amylopectin inthe growth rings, and the lamellar architecture of the crystallinelayers within these rings, are still not fully understood. Nor is thedistribution and role of water, which acts as a plasticiser and influ-ences the crystallinity of the structure (Donald, 2001; Perry &Donald, 2000). Granules contain about 10% water, which is probablynot distributed uniformly. The concentric pattern of semi-crystallinelayers is responsible for the birefringence of native starch granuleswhen viewed under polarizing light microscopy.

Amylopectin chains with more than 10 glucose units are orga-nized into double helices, which are arranged into either A- orB-crystalline forms that may be identified by characteristic X-raydiffraction (XRD) spectral patterns. The double helical structureswithin the A- and B-type crystalline forms are essentially the same(Gidley, 1987; Imberty, Buleon, Tran, & Perez, 1991), but the packingof the helices in the A-type crystalline structure is more compactthan in B-type crystallites, which have a more open structure witha hydrated core. The A-type crystal pattern has amylopectinmolecules with shorter chains (Jane, 2006). Cereal starches tend tohave the A-type pattern, whereas tuber starches and amylose-richstarches yield the B-type pattern, although both types may occurtogether. Legume, root and some fruit and stem starches yield anintermediate C-type pattern, but whether this is a mixture of theA- and B-type patterns or a distinct form is not clear (Buleon et al.,1998; Cairns, Bogracheva, Ring, Hedley, & Morris, 1997).

Starches from wheat (Triticum aestivum L.), barley, rye and triti-cale have a bimodal granule size distribution. In wheat, there is onepopulation of small spherical granules ranging in size fromapproximately 1 to 10 mm, which are referred to as B-granules, andanother population of larger lenticular-shaped granules rangingfrom about 15 to 40 mm, known as A-granules (Peng, Gao, Abdel-Aal,Hucl, & Chibbar, 1999; Shinde, Nelson, & Huber, 2003; Soulaka &Morrison, 1985; Tang, Ando, Watanabe, Tajeda, & Mitsunaga, 2001).The biosynthesis of A- and B-granules differs during grain filling,resulting in differences in the molecular organization of the amyloseand amylopectin fractions and molecular architecture of theamylopectin (Bechtel, Zayas, Kaleikau, & Pomeranz, 1990; Parker,1985; Shinde et al., 2003). The A-granules make up a much greaterproportion of wheat starch by weight but are much fewer in numberthan B-granules. The A- and B-granules differ in structural andfunctional properties, including amylopectin of B-granules havingmore short chains and fewer medium and long chains than that of A-granules (Ao & Jane, 2007; Liu, Gu, Donner, Tetlow, & Emes, 2007;Peng et al., 1999; Salman et al., 2009; Shinde et al., 2003; Tang et al.,2001; Vermeylen, Goderis, Reynaers, & Delcour, 2005).

4. Analyzing starch structure

The structure of amylose and amylopectin, and the form andcrystallinity of starch granules, have been studied extensively usingmany complementary approaches. Amylose helices form inclusioncomplexes with polyiodide giving a blue colour, which has longbeen used as a colorimetric test for starch. Although rapid andconvenient, this method is subject to inaccuracies because thewavelength of maximum absorbance of amylose–iodine complexesvaries with DP, and the formation of amylopectin–iodinecomplexes that can also absorb at interfering wavelengths (Gibson,Solah, & McCleary, 1997). Separation of amylose from amylopectinby size exclusion chromatography or by using the lectin conca-navalin A are alternative approaches to amylose measurement instarches but are less suited to high throughput analyses (Gibsonet al., 1997). The fine structure of amylopectin has been examinedusing highly purified amylolytic enzymes (Manners, 1989).

A combination of techniques have been used to study themolecular organization within starch granules, to obtain estimatesof the thickness of the crystalline regions, and explore how amyloseaffects amylopectin clusters in native starch granules (for example:Bertoft, 2004; Blanshard, Bates, Muhr, Worcester, & Higgins, 1984;Donald et al., 2001; Kozlov et al., 2006; Lopez-Rubio, Htoon, &Gilbert, 2007; Vermeylen et al., 2006; Waigh, Perry, Riekel, Gidley,& Donald, 1998). Small-angle scattering techniques measuredifferences in electron density distribution, whereas diffractiontechniques are indicative of crystallinity of the material. Differentialscanning calorimetry (DSC) identifies melting and crystallizationevents as well as glass transition temperatures. Imaging techniquessuch as light microscopy, scanning and transmission electronmicroscopy, scanning probe techniques such as atomic forcemicroscopy (AFM), spectroscopic methods such as nuclearmagnetic resonance and Fourier transformed infra-red spectros-copy are other approaches that have been used to obtain structuralinformation on starch granules and molecules (Dang, Braet, &Copeland, 2006; Dang & Copeland, 2003; Gunning et al., 2003; Liu,Guo, Hu, & Li, 2001; McIntire & Brant, 1999; Szymonska & Krok,2003).

These analytical methods provide explicit information on themolecular constituents and their organization in starch granules. Incomparison, functionality is usually assessed with tests that arerelevant to a particular application and are based on measuring thecollective properties of a material. Relating an observed functionaleffect to a physicochemical change in a specific molecular constit-uent is often difficult. The bridging of this gap is considered in thenext sections.

5. Functional properties of starch

Most starch consumed by humans has undergone some form ofprocessing, which usually involves heating in the presence ofmoisture under shear, and then cooling. During heat treatment, thestarch granules are gelatinized, losing their crystallinity andstructural organization. On cooling, the disaggregated starchmolecules first form a gel and then retrograde gradually into semi-crystalline aggregates that differ in form from the native granules.Starch-rich foods that have been cooked and cooled often containsubstantial amounts of retrograded starch. Understanding the stepsthat occur during gelatinization and retrogradation of a particularstarch are key steps to better predicting the functional properties ofprocessed starch from knowledge of the structure of nativegranules.

Gelatinization occurs when native starch is heated in the pres-ence of sufficient moisture. The granules absorb water and swell,and the crystalline organization is irreversibly disrupted. Accordingto the theory of Jenkins and Donald (1998), water first enters the

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amorphous growth rings, and at a certain degree of swelling,disruptive stress is transmitted through connecting molecules fromthe amorphous to the crystalline regions. Amylose molecules beginto leach from the granules as they are disrupted under shear andthe viscosity of the resulting paste increases to a maximum, whichcorresponds to the point when the number of swollen but stillintact starch granules is at a maximum. The maximum is followedby a decrease in paste viscosity, as the granules rupture and starchmolecules are dispersed in the aqueous phase. The rate and extentof swelling and breakdown are dependent on the type and amountof starch, the temperature gradient, shear force and the composi-tion of the mixture, for example the presence of lipids and proteins(Debet & Gidley, 2007). The gelatinization temperature of moststarches is between 60 and 80 �C. In general, there is a negativerelationship between the amylose content of starch and the gela-tinization temperature and peak viscosity.

As the starch paste cools, the viscosity increases due to theformation of a gel held together by intermolecular interactionsinvolving amylose and amylopectin molecules. In gels that containabout 25% amylose, the starch molecules form a network resultingin a firm gel, in contrast to waxy starch gels, which are soft andcontain aggregates but no network (Tang & Copeland, 2007b). Onstanding, starch gels retrograde and form insoluble B-type crys-tallites due to association of linear regions of a-[1 / 4] linkedglucose units in the polymers (Gidley, 1989). Retrogradation is anongoing process occurring over an extended period. Amyloseretrogrades over minutes to hours and amylopectin over hours todays, depending on the ability of the branched chains to formassociations. The retrogradation of amylose in processed foods isconsidered to be important for properties relating to stickiness,ability to absorb water, and digestibility, whereas retrogradation ofamylopectin is probably a more important determinant in thestaling of bread and cakes. In rice, amylose contributes texture andstickiness, whereas gelatinization temperature, cooking andpasting properties are more closely related to amylopectin (Tran,Okadome, Murata, Homma, & Ohtsubo, 2001).

Gelatinization behaviour and the pasting profiles of flour–waterand starch–water mixtures are commonly monitored using a RapidVisco Analyser (RVA), which is a heating and cooling viscometerthat measures the resistance of a sample to controlled shear. TheRVA is considered to simulate food processing and is used to relatefunctionality to structural properties, as discussed in the compre-hensive monograph edited by Crosbie and Ross (2007). A typicalRVA profile of starch gelatinization from the authors’ laboratory(Fig. 1) shows the viscosity increase to a maximum, followed bya decrease to a minimum value as the granules rupture (referred to

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Fig. 1. Typical RVA profile of rice starch.

as the breakdown). As the temperature is decreased, the viscosityagain increases from the minimum to a final value, which is referredto as the setback. The peak time and peak viscosity are indicative ofthe water-binding capacity of the starch and the ease with whichthe starch granules are disintegrated, whereas higher setbackvalues are usually correlated with the amylose content of the starch.The RVA provides a convenient way of studying the effects ofadditives on rheology of starch systems (Deffenbaugh, Lincoln, &Walker, 1989; Ravi, Sai Manohar, & Haridas Rao, 1999; Tang &Copeland, 2007a). The RVA parameters have been correlated withtexture and product quality. For example, larger breakdown value isconsidered to be an indicator of better palatability of cooked rice(Tran et al., 2001). Viscometric analyses can detect differences infunctional properties between starches and flours from differentvarieties or growth environments that may not be evident fromconventional chemical analyses (Dang & Copeland, 2004).

6. Starch–lipid complexes

Complexes between amylose and lipids, such as fatty acids,lysophospholipids and monoacylglycerides, can significantlymodify the properties and functionality of starch in ways that are ofinterest to the food industry and for human nutrition. For example,complexation with lipids reduces the solubility of starch in water,alters the rheological properties of pastes, decreases swellingcapacity, increases gelatinisation temperature, reduces gel rigidity,retards retrogradation and reduces the susceptibility to enzymichydrolysis (Biliaderis & Seneviratne, 1990; Crowe, Seligman, &Copeland, 2000; Guraya, Kadan, & Champagne, 1997; Holm et al.,1983; Karkalas & Raphaelides, 1986; Kaur & Singh, 2000; Nierle & ElBaya, 1990; Ozcan & Jackson, 2002; Tufvesson, Wahlgren, & Eli-asson, 2003a, 2003b). A quick scan of the patent literature indicatesthat starch–lipid complexes have long been used in numerousapplications in the food industry, to reduce stickiness of starchyfoods, improve freeze–thaw stability, to retard staling in bread andbiscuits, and as dough conditioners and crumb softeners in breads.Starch–lipid compositions are also used as fat replacers, stabilizersand thickener in foods. Complexation of starch with endogenous oradded lipids may occur during gelatinization, or when starch andlipids are heated together in the RVA. A variety of cooking andextrusion processes have been used to form complexes underconditions that mimic processing (Bhatnagar & Hanna, 1994; Fanta,Shogren, & Salch, 1999). Starch–lipid complexes have been studiedextensively using the same techniques as for analysis of starchgranules and starch molecules, namely DSC, XRD, iodine-bindingcapacity, viscometry, and image analysis. The complexes may bequantified using competitive iodine-binding studies that measurethe reduction in iodine-binding colour when complexes are formed(Kaur & Singh, 2000, Tang & Copeland, 2007a).

The glucosyl hydroxyl groups of a[1 / 4] glucose chains arelocated on the outer surface of the helix. The inner core is a morehydrophobic cavity lined with methylene groups and glycosidicoxygens, and can form inclusion complexes with a variety oforganic and inorganic ligands, for example, dimethyl sulfoxide,potassium hydroxide, polyiodide ions, lipids and linear alcohols(Immel & Lichtenthaler, 2000; Nuessli, Putaux, Le Bail, & Buleon,2003). These complexes form single helices that are generallyinsoluble in water and have a V crystalline form with a character-istic XRD pattern different from the A- and B-crystal forms. Theinner diameter of the helix (six, seven or eight glucose residues-per-turn) is controlled by the nature of the complexing agent(Biliaderis & Galloway, 1989; Helbert & Chanzy, 1994).

Inclusion complexes with lipids form mainly with the amylosecomponent of the starch, and hence the amylose to amylopectinratio is an important factor that produces variability in the ability ofnatural starches to bind lipids. Lipid-complexed amylose has been

L. Copeland et al. / Food Hydrocolloids 23 (2009) 1527–1534 1531

confirmed to occur in native cereal starch granules, but as cerealstarches contain about 1% lipid, only 15–55% of the amylose fractionis complexed with lipid (Morrison, 1988, 1995). Due to its highdegree of branching, the lipid binding capability of amylopectin isconsidered to be much weaker than amylose. There is little directevidence to suggest that amylopectin forms true inclusioncomplexes with lipids, although some studies have led to proposalsthat favour interaction of some lipids and surfactants with outeramylopectin branches (Eliasson, 1994; Hahn & Hood, 1987; Vill-wock, Eliasson, Silverio, & BeMiller, 1999).

The extent of crystallinity and thermal stability of complexesbetween amylose and complex-forming lipids increase withincreasing amylose chain length (Gelders, Duyck, Goesaert, &Delcour, 2005; Godet, Bizot, & Buleon, 1995a). The effect of the lipidcomponent on complex formation is less well defined. The heatstability of complexes between fatty acid anions and amyloseincreases with fatty acid chain length (Godet, Tran, Colonna,Buleon, & Pezolet, 1995b; Tufvesson et al., 2003a, 2003b), anddecreases with unsaturation (Hahn & Hood, 1987). Monoglyceridesof saturated fatty acids with chain lengths from C12 to C18 are veryeffective amylose complexing agents in aqueous solution at 60 �C,as are trans-unsaturated analogues, whereas the cis-unsaturatedanalogues form thermally less stable complexes (Eliasson & Krog,1985; Morrison, 1988; Ozcan & Jackson, 2002; Raphaelides & Kar-kalas, 1988; Riisom, Krog, & Eriksen, 1984). Maximal complexformation occurs at a different concentration for different lipids,depending on water solubility and critical micellar concentration ofthe lipid (Tang & Copeland, 2007a). Above a certain concentration,some lipids (for example, palmitic and stearic acids) tend to self-associate in preference to forming starch–lipid complexes, whichhelps to explain why a particular lipid has been described asforming strong complexes with amylose in some studies and weakor no complexes in others. There is no evidence to indicate thatdiglycerides or triglycerides form inclusion complexes withamylose (Morrison, 1988, Tang & Copeland, 2007a, 2007b).

Studies with NMR and molecular modelling of amylose–lipidcomplexes indicate that the hydrophobic fatty acid chain isincluded inside the amylose helix, whereas the polar groups of fattyacids and monoacylglycerols are too bulky to enter the helical tube(Buleon et al., 1998; Nimz, Gessler, Uson, Sheldrick, & Saenger,2004; Snape, Morrison, Maroto-Valer, Karkalas, & Pethrick, 1998).On this basis, amylose–lipid complexes have been represented bya model in which the aliphatic moiety of a lipid (for example, a fattyacid or monoacyglycerol) may be inserted at each end of anamylose chain, as represented diagrammatically in Fig. 2.

The effect of lipids on viscosity of starch pastes has beenmonitored by viscometry, and the RVA has been used as a simpleand expedient method to mix starch and lipids and quantifycomplex formation (Tang & Copeland, 2007a). Compared withgranular starch or defatted starch, a greater final viscosity and totalsetback are characteristics for starch mixed with fatty acids andmonoglycerides, and peak viscosity may also decrease (Deffen-baugh, Lincoln, & Walker, 1990; Karkalas & Raphaelides, 1986;Kaur & Singh, 2000; Liang, King, & Shih, 2002; Ozcan & Jackson,2002; Ravi et al., 1999). The increased setback of starch in thepresence of lipids has been explained by the formation of gels with

Fig. 2. Schematic representation of a complex of

more widely spaced junction zones due to the reduced availabilityof amylose to form networked aggregates. The networks observedwith AFM in starch-only gels were absent when starch–lipidcomplexes were formed (Tang & Copeland, 2007b).

Two thermally distinct forms of amylose–lipid complexes havebeen identified: an amorphous structure (termed Form I) with anendothermic transition in the DSC near 100 �C, and crystallinestructures with DSC transitions at about 115 �C (Form IIa) and125 �C (Form IIb). Form I appears to have a random distribution ofaggregated helices, whereas Form II has a crystalline organizationof amylose complexes. Form II requires higher temperatures andlonger time to form, and is responsible for the distinctive V-typeXRD pattern (Biliaderis & Galloway, 1989; Gelders et al., 2005;Godet, Bouchet, Colonna, Gallant, & Buleon, 1996; Seneviratne &Biliaderis, 1991; Tufvesson & Eliasson, 2000). The polar head of thefatty acid seems to be an important factor in allowing amylosechains to interact and for Form II complexes to occur; they hardlyarise with shorter fatty acids (C10 and C12), in contrast to thecorresponding monoglycerides. This has been proposed to be dueto steric and electrostatic repulsion of the carboxyl group of thefatty acid, which is outside the amylose helix (Tufvesson et al.,2003a, 2003b). Both Forms I and II of amylose–lipid complexes maybe present in processed starch-based foods depending on themethod of processing. Protein also has an effect on the properties ofstarch–lipid complexes. Such multiple interactions are likely to becommon in cooked starchy foods and may influence their func-tionality, but studies are limited (Zhang & Hamaker, 2003, 2004).

7. Enzymic digestion of starch

A functional property of starch that is of particular interest fornutrition is its susceptibility to digestion. Starch that is notdegraded rapidly by human digestive enzymes in the upper gut hasbeen associated with health benefits due to a slower release ofglucose into the blood stream resulting in reduced postprandialglycemic and insulin responses. The name ‘‘resistant starch’’ is usedto describe starch in foods that is incompletely digested. Assummarized by Topping and Clifton (2001), the resistance of starchto digestion may be due to its intrinsic properties, the extent towhich food is chewed, the result of changes during processing,retrogradation, chemical modification, or due to interactions withother food constituents, especially lipids. Undigested or partiallydigested starch that passes from the upper part of the gut into thelower gut is considered to be a prebiotic, in that it is a goodsubstrate for beneficial gut microflora associated with colonichealth. Trying to obtain starches with slow-digesting properties isan important objective for the food industry.

The digestion of starch has been the subject of many inves-tigations, mostly involving in vitro measurement of the suscepti-bility of starches to attack by different enzymes, rather thanmeasuring actual digestibility in vivo. Depolymerization of starchin the upper gut is effected by a-amylases, which cleave thea-[1 / 4] links, and amyloglucosidases and isoamylases, whichbreak a-[1 / 6] glucosidic bonds. The rate and extent of amylolytichydrolysis of granular starches varies according to botanical origin.However after cooking, differences in susceptibility of starch to

amylose with two monopalmitin molecules.

Fig. 3. Partially degraded granules of waxy (left) and a high-amylose (right) wheat starch after 2 h with pancreatic alpha-amylase (J. Blazek et al., 2009).

L. Copeland et al. / Food Hydrocolloids 23 (2009) 1527–15341532

enzymic attack are related more to the products of gelatinizationand, perhaps more critically, retrogradation.

Granule characteristics considered to influence susceptibility toattack by a-amylase include: crystallinity, granule size and avail-able specific surface, amylose to amylopectin ratio, porosity,structural inhomogeneities and degree of integrity (Kong, Kim,Kim, & Kim, 2003; Planchot, Colonna, Buleon, & Gallant, 1997; Ring,Gee, Whittam, Orford, & Johnson, 1988). A-type crystals tend to bemore resistant to enzymic digestion than the B-crystal form. Inhi-bition of a-amylase by hydrolysis products such as maltose andmaltotriose may also be relevant (Colonna, Buleon, & Lemarie,1988; Leloup, Colonna, & Ring, 1991). Crystalline lamellae are lessreadily attacked by enzymes than amorphous regions. Gelatinizedstarches are digested much more rapidly than native starch gran-ules and retrograded starch, consistent with the notion that accessof enzymes to the starch molecules may be the key factor that limitsenzymic hydrolysis. Diffusion effects on the release of productsfrom granules may also be significant.

The kinetics of a-amylolysis of intact granules are characterisedby an initial rapid hydrolysis phase followed by a slower phase(Colonna et al., 1988). Waxy starch granules are attacked morereadily than starch granules with significant amounts of amylose(Jane, 2007). Water absorption, and presumably enzyme access,first occurs in the amorphous regions of the granules. Therefore, inwaxy starch water absorption may be less impeded than in starcheswith amylose in the amorphous layers. Attack by a-amylase onwheat starch is characterised by the formation of holes and pref-erential disruption of the core of the granule (Gallant, Derrien,Aumaitre, & Guilbot, 1973). Initially, the granular surface is attackedand susceptible zones become pitted. These pits enlarge andnumerous channels are formed towards the centre of the granule.SEM observations reveal major differences in the appearance ofpartially degraded starch granules of different composition (Fig. 3).Starch granules from some sources seem to be corroded uniformlyin vitro by added enzymes, whereas others contain a mixture ofhighly damaged and relatively undamaged granules, as a result ofan ‘‘all-or-none’’ mode of attack (Kitahara, Suganuma, & Nagahama,1994).

Both added and endogenous lipids have an influence on starchdigestibility and resistant starch formation (Crowe et al., 2000;Gelders et al., 2005; Guraya et al., 1997; Holm et al., 1983; Nebesny,Rosicka, & Tkaczyk, 2002; Szczodrak & Pomeranz, 1991). Complexeswith greater crystallinity are more resistant to enzymatic degra-dation. The rate and extent of degradation by bacterial andpancreatic amylases has been assessed to decrease in the order:Form I> Form IIa> Form IIb (Tufvesson et al., 2003a). Complexeswith long chain saturated monoglycerides are more resistant toenzymic breakdown than complexes with shorter aliphatic chains

or greater degrees of unsaturation of the ligand (Biliaderis & Sen-eviratne, 1990, Tufvesson, Skrabanja, Bjorck, Liljeberg Elmståhl, &Eliasson, 2001).

8. Concluding comments

The main source of starch in our diet is from cereal grains, whichare unpalatable and largely indigestible in their raw state. Pro-cessing of grains provides foods that appeal to consumers in taste,nutritional value, and shelf life. Our ability to relate starch granulestructure to suitability for a particular food manufacturing processor its nutritional qualities depends not only on knowledge of thegenetic and environmental factors that control starch biosynthesis,and in turn granule morphology, but also on how the material isprocessed. There has been considerable progress towards relatingthe morphological complexity of native starch granules to pro-cessing properties, although there are still many gaps in ourknowledge. Amylose influences the packing of amylopectin intocrystallites and the organization of the crystalline lamellae withingranules. A better understanding of the location of amylose instarch granules may improve our ability to relate structure toproperties that involve water absorption, for example, swelling,gelatinization, and susceptibility to enzymic attack. Thermalproperties and gel formation appear to be influenced by bothamylose content and amylopectin architecture. While there is someunderstanding of the influence of genotype on starch structure andgranule morphology, our knowledge of how these are affected byenvironmental factors during crop growth is still limited. Recentresearch by the authors using a genetically diverse population ofwheat varieties (Blazek et al., 2009; Salman et al., 2009) has shownthat starches may be similar chemically, but can have significantlydifferent functional properties. Thus, while amylose content islikely to have an important bearing on the functional properties ofstarch, subtle structural variations in the molecular architecture ofamylopectin introduces uncertainty into the prediction of func-tional properties from amylose content alone. Processing destroysthe native structure of granules and creates new molecular inter-actions, thereby greatly complicating the relationship betweenform and function. This may present particular difficulties togaining an understanding of the nutrition functionality of starch,since much of the starch we consume has lost its highly organizedstructure and granular properties during processing.

Acknowledgement

JB, HS and MT were supported by scholarships from the ValueAdded Wheat Cooperative Research Centre Ltd.

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