Richard F. Tester

Starch structure and digestibility Enzyme-Substrate relationship

Digestion of starch is effected by hydrolysing enzymes in a complex process which depends on many factors; these include the botanical origin of starch, whether the starch is amorphous or crystalline, the source of enzymes, substrate and enzyme concentration, temperature and time, as well as the presence of other substances in the multicomponent matrix in which starch occurs naturally, e.g. cereal grains. Native starch is digested (i.e. hydrolysed) slowly compared with processed (gelatinised) starch whose crystallinity has been lost and where the accessibility of substrate to enzymes is greater and not restricted by α-glucan associations such as double helices (especially in crystallites) or amylose-lipid complexes (in cereal starches). The restriction of starch digestion (primarily in the human digestive system) due to forms which are resistant to hydrolysis has led to the concept of dietary ‘resistant-starch’. Different forms of resistance can be identified which hinder hydrolysis. With regard to digestibility, whether in the human or animal digestive tract, it is important to understand the mechanisms of enzymatic hydrolysis, and the consequence of incomplete digestion i.e. the potential loss of glucose as a valuable source of energy. This review deals with starch hydrolysis by specific amylases in model (in vitro) and more broadly in in vivo systems. The optimisation of starch hydrolysis and digestion is discussed in the light of modern knowledge.

The effects of non-starch polysaccharides on the extent of gelatinisation, swelling and α-amylase hydrolysis of maize and wheat starches.

Starch hydrolysis (α-amylase) and gelatinisation characteristics were evaluated in the presence of five different polysaccharides (arabic, carrageenan, guar, pectin and xanthan) with different starch to solution ratios and polysaccharide concentrations therein. Using relatively high starch/solution ratios (1:10) at concentrations (<5%) of the polysaccharides that permitted some starch mobility (in the solutions) it was possible to restrict swelling of starch granules and consequently restrict the amount of starch gelatinisation. With lower solution ratios (1:0.8, 1:1.7, 1:3.3 and 1:5.0, especially 1:0.8 and 1:1.7) it was possible to model the effects of the polysaccharides in terms of displacing water and restricting gelatinisation with respect to the (modified) Flory–Huggins equation. With and without taking into account specific hydration of the polysaccharides, it was possible to show a linear relationship between the end point of gelatinisation (Tc) and the volume fraction of water in the system (v1). The implications in terms of product processing have been discussed.

Annealing of maize starch

Abstract A comparison was made between the annealing characteristics of commercial waxy, normal and amylomaize starches. The waxy and normal starches responded similarly to annealing where improved registration of amylopectin double helices is proposed, with perhaps optimisation of hydrogen bonding along the full length of individual helices. This is attributed to similar amylopectin crystalline structure of these starches. It is considered unlikely that the free amylose (FAM) forms any significant numbers of amylose–amylose or amylopectin–amylose double helices throughout its length, but may randomly form some double helical regions in the crystalline shells created essentially by the amylopectin. Single amylose helices are also considered unlikely. Probably lipid complexed amylose (LAM) is excluded from the amylopectin crystallites and is primarily located in amorphous shell regions (between crystalline shells) and consequently affects hydration (and glass transition, Tg) of these regions. In amylomaize starches, however, the larger amylopectin chain lengths, broad gelatinisation endotherm, leaching characteristics and double helix content (by NMR) indicates that FAM does here play a significant part in the crystalline regions of the starch. Annealing of amylomaize crystallites probably reflects enhanced registration (and possibly compartmentalisation) of amylopectin–amylopectin, amylopectin–amylose and amylose–amylose helices.

The effects of ambient temperature during the grain-filling period on the composition and properties of starch from four barley genotypes

Starch was isolated from four genotypes of barley (one waxy, two normal and one high-amylose) grown at constant ambient temperatures of 10, 15, and 20 °C. There was evidence of physiological stress in grain grown at the higher temperatures, notably reduced starch accumulation, smaller A- and B-granules and fewer B-granules. In the waxy and normal genotypes amylose (AM) and amylopectin (AP) contents were little affected by increasing ambient temperature, but the lipid contents of the starches did show a strong response. No differences in the fine structure of AP in response to temperature were detected. Starch gelatinization temperatures (GT) were 50–55 °C, 52–62 °C and 60–63 °C in starches from grain grown at 10, 15 and 20 °C, respectively. Since the native starches could all be annealed to give similar elevated GT values (72–73 °C, the range of GT in the native starches was probably due to differences in the perfection of AP crystallinity which increased with barley growth temperature. All starches showed marked differences in their swelling curves which clearly related to ambient temperature. Swelling began at the same temperature as gelatinization (loss of crystalline order), At 80 °C, when there was no detectable order, swelling was determined primarily by AP content (45·3–97·6% in the 12 starches) but it was markedly inhibited by lipid (a temperature-dependent variable) probably through a mechanism which involved formation of AM-lipid inclusion complexes.

Influence of growth conditions on barley starch properties

Air equilibrated barley starch comprises amylopectin, amylose, lipid and water. The structure of amylose and amylopectin, and the proportion of amylose in granules is under genetic control and is therefore subject to genotypic variation. The amount of lipid (which is essentially all lysophospholipid) is similarly under genetic control. Environment and especially environmental temperature do, however, have a regulatory effect on the size of starch granules, the amylose to amylopectin ratio and the amount of lipid (which is essentially all complexed with amylose) within barley starch. High growth temperatures probably facilitate amylopectin crystallisation and increase gelatinisation temperatures, (and to some extent the enthalpy of gelatinisation), but delay the onset and depress the extent of swelling of granules when heated in water.

Annealing of wheat starch

Abstract The moisture, time and temperature dependence of annealing for one commercial and 10 laboratory extracted starches (from five soft and five hard wheats grown in various places in England in 1994) were investigated. Annealing was found to occur at least 15°C below the onset gelatinisation temperature (T o ), the extent of which was time dependent, but was much more evident (as indicated by the relative increase in gelatinisation temperatures) the closer to T o the annealing temperature was set. Annealing could be initiated when the starch contained 20% by weight moisture, but the process was restricted unless the moisture content exceeded 60%. 13 C-CP/MAS-NMR indicated that the number of double helices remained constant post-annealing and it is proposed that annealing improves the crystalline register of double helices, thereby ‘perfecting’ starch crystallites rather than promoting the formation of additional double helices. This perfection of crystallites is possibly initiated by incipient swelling and the resulting mobility of amorphous α-glucans which facilitates ordering of double helices and, probably, greater ordering of the amorphous regions themselves.

Resistance to acid hydrolysis of lipid-complexed amylose and lipid-free amylose in lintnerised waxy and non-waxy barley starches

Abstract Waxy barley starches (0.8–4.0% lipid-complexed amylose = L·AM, 0.9–3.4% lipid-free amylose = F·AM) and non-waxy barley starches (6.1–7.2% L·AM, 23.1–25.9% F·AM) were lintnerised by steeping in 2 M HCl at 35°C for 140 h. Material solubilised from the waxy starches was estimated to be 70.7% of their amylopectin (AP) plus 3.7% of their L·AM and F·AM, and material solubilised from the non-waxy starches was estimated to be 70.7% of their AP plus 28.9% of their L·AM and F·AM. The polysaccharide components of the insoluble residue were characterised by HPLC, GPC, and λ max of the polyiodide complex. I was concluded that short chain-length (CL 16) material was from external chains of AP, intermediate material (modal CL 46) was from retrogated F·AM, and longer chain residues (CL 77, 120–130_ were from lipid-complexed segments of L·AM. The starch lysophospholipids were completely hydrolysed to free fatty acids which remained complexed with L·AM residues. This was shown by the 13 C CP/MAS-NMR spectrum which had a clear resonance at 31 ppm from mid-chin methylene carbons of fatty acids in complexes. The C-1 signal of the L·AM residues also included a feature at 104 ppm indicative of single V 6 AM helices. The wide-angle X-ray diffraction patterns of the residues of non-waxy starches were Cc-type ( = mixed A + B types), whereas the spectra of the original starches were A-type. It is suggested that, during the early stages of lintnerisation, amorphous (F·AM was partially hydrolysed into material (CL 13 C CP/MAS-NMR spectra, which were consistent with a mixture of double helices and V-type glycosidic conformations, with only a small proportion of non-ordered regions. Broad DSC endotherms were found for both waxy (50–110°C) and non-waxy (50–110°C) lintner residues, which were assigned to disordering of double helices from short chains (modal CL 16) for waxy residues, together with disordering of longer chains (modal CL 46) in double-helix residues of F·AM and also V-helix residues of L·AM for non-waxy starch residues.

In vitro binding of calcium, iron and zinc by non-starch polysaccharides

The in vitro binding capacity of eight non-starch polysaccharides (agar, κ-carrageenan, gum xanthan, gum arabic, gum karaya, gum tragacanth, pectin and gum guar) was measured by equilibrium dialysis in neutral and acidic (0.1M HCl) solutions in the presence of divalent cations (Ca2+, Zn2+). No significant binding was observed in acidic conditions while, in neutral solutions, the extent of binding was correlated (P<0.1) to the cation-exchange capacity of the polysaccharides. It is apparent that the interactions are essentially electrostatic in nature, due to the presence of ionised carboxyl (uronic/pyruvic acids) and sulphated groups, in polyanionic polysaccharides. By contrast, significant binding occurs with Fe3+ in acidic conditions, presumably due to complexation (chelation). These data provide a clear insight into how non-starch polysaccharides interact with minerals and the potential nutritional consequence in terms of bioavailability.

Occurrence of aflatoxin M1 in randomly selected North African milk and cheese samples

Forty-nine samples of raw cows milk and 20 samples of fresh white soft cheese were collected directly from 20 local dairy factories in the north-west of Libya and analysed for the presence of aflatoxin M1 (AFM1). The samples were analysed using a high-performance liquid chromatography technique for toxin detection and quantification. Thirty-five of the 49 milk samples (71.4%) showed AFM1 levels between 0.03 and 3.13 ng ml−1 milk. Multiple analyses of five milk samples free of AFM1 artificially contaminated with concentrations of AFM1 at 0.01, 0.05, 0.1, 1.0 and 3.0 ng ml−1 showed average recoveries of 66.85, 72.41, 83.29, 97.94 and 98.25%, with coefficients of variations of 3.77, 4.11, 1.57, 1.29 and 0.54%, respectively. Fifteen of 20 white soft cheese samples (75.0%) showed the presence of AFM1 in concentrations between 0.11 and 0.52 ng g−1 of cheese. Multiple assays of five cheese samples free of AFM1 spiked with different concentration of AFM1 (0.1, 0.5, 1.0 and 3.0 ng g−1) showed average recoveries of 63.23, 78.14, 83.29 and 88.68%, with coefficients of variation of 1.53, 9.90, 4.87 and 3.79%, respectively. The concentrations of AFM1 were lower in the cheese products than in the raw milk samples.

Properties of damaged starch granules: composition and swelling properties of maize, rice, pea and potato starch fractions in water at various temperatures

A range of cereal and non-cereal starches (maize, waxy maize, rice, waxy rice, pea and potato) were ball milled to obtain various levels of damage. The susceptibility of the different starches to ball milling-induced damage was related to granule composition, dimensions, size distribution, gelatinization parameters by differential scanning calorimetry, swelling factors determined by dye exclusion, and leaching characteristics of α-glucan from the granules heated in water. The formation of granule remnants, gel-forming material and soluble amylopectin fragments from the damaged starches was investigated and related to gel volume formed at different temperatures.