High-amylose Wheat Research Articles

Milling and rheological properties of high amylose wheat

The technological quality of a high amylose bread wheat variety (amylose/amylopectin 1:0.7) was explored performing different milling protocols, with subsequent evaluation of the chemical composition and rheological properties of the milling fractions and recombined flours. High amylose wheat yielded a lower amount of flour (<50%) and less refined products compared to control line. Grain debranning improved flour yields, releasing more refined reduction products. The milling of high amylose wheat with a hard-wheat diagram enhanced flour yield and refining rate of products, also giving a semolina type flour that can be employed e.g. for the production of pasta and couscous. The high amylose flours are characterized by relevant levels of resistant starch (27% of total starch on average) that make them suitable for the production of functional foods in agreement with the EU Regulation 432/2012. In addition, from the experimental results, it was put in evidence that official protocols for rheological analysis of flours (gluten index, alveograph and falling number) should be adapted for high amylose wheat samples, due to the impact that the different starch composition has on dough properties, specifically on water absorption and viscosity. Overall, rheological features of high amylose wheat flours exhibited higher water absorption, stability, and altered starch pasting properties.

High-amylose wheat bread with reduced in vitro digestion rate and enhanced resistant starch content

High amylose wheat flour (HAWF) is a relatively new food ingredient, with potential for enhanced nutritional functionality of slower digestion and greater resistant starch levels compared to wild-type wheat flour (WTWF) products. This study focusses on the role of amylose content in the baking process and eating qualities of bread, made using an industry-standard rapid dough method. DSC, XRD and polarized light microscopy were used to monitor changes of ordered structure in starch after baking, and showed more retained crystalline structure in the high-amylose wheat bread (HAWB) than wild-type wheat bread (WTWB). Moreover, HAWB had lower loaf volume, more dense crumb structure, as well as higher hardness than WTWB. These starch structural and local density features subsequently lead to bread made with 100% or 50% HAWF being more resistant to enzyme hydrolysis compared to WTWB. Resistant starch levels were significantly increased by ∼11 and 12 times in bread made using 100% HAWF with amylose content 71% and 84% respectively, and ∼6 times in bread prepared with 50% incorporation HAWF (84% AM) than WTWB. The nutritional and physical properties of bread are highly correlated to starch amylose content (AM), starch thermal properties and crumb hardness, with AM playing the leading role. This study identifies the structural basis for flour amylose content effects in baking and nutritional functionality and can help guide development of nutritionally-enhanced foods with HAWF.

Molecular-structure evolution during in vitro fermentation of granular high-amylose wheat starch is different to in vitro digestion

This study investigates the evolution of the distributions of whole molecular size and of chain length of granular wheat starches (37 ~ 93% amylose content), subjected to in vitro fermentation with a porcine faecal inoculum or digestion with pancreatic enzymes. The results showed that the molecular structures of high-amylose starch (HAS) unfermented residues largely remained unchanged during the fermentation process, while wild-type starch (37% amylose content) showed a preferential degradation of the amylopectin fraction. In contrast, under simulated digestion conditions, the undigested residues of HAS showed structural changes, including a decrease in amylose content, a shift of amylose peak position towards lower degrees of polymerisation, and an enzyme-resistant fraction. These changes of starch structure are likely to be dependent on the different starch-degrading enzyme activities present in pancreatic vs. microbial systems. Molecular changes in response to fermentation metabolism revealed by size-exclusion chromatography can help understand the microbial utilization of resistant starch.

A High Amylose Wheat Diet Improves Gastrointestinal Health Parameters and Gut Microbiota in Male and Female Mice.

High amylose wheat (HAW) contains more resistant starch than standard amylose wheat (SAW) and may have beneficial effects on gastrointestinal health. However, it is currently unclear whether these effects differ according to the level of HAW included in the diet or between males and females. Male and female C57BL/6 mice (n = 8/group/sex) were fed SAW65 (65% SAW; control), HAW35 (35% HAW), HAW50 (50% HAW) or HAW65 (65% HAW) diet for eight weeks. Female but not male, mice consuming any amount of HAW exhibited accelerated gastric emptying compared to SAW65 group. In both sexes, relative colon weights were higher in the HAW65 group compared to SAW65 group and in females, relative weights of the small intestine and cecum were also higher in the HAW65 group. In females only, colonic expression of Pyy and Ocln mRNAs were higher in the HAW65 group compared to HAW35 and HAW50 groups. In both sexes, mice consuming higher amounts of HAW (HAW50 or HAW65) had increased fecal bacterial load and relative abundance of Bacteroidetes phylum and reduced relative abundance of Firmicutes compared to SAW65 group. These data are consistent with a beneficial impact of HAW on gastrointestinal health and indicate dose-dependent and sex-specific effects of HAW consumption.

Evaluation of an Ileorectostomised Rat Model for Resistant Starch Determination.

The human ileostomy model, widely considered the benchmark for determining in vivo starch digestibility, has disadvantages. The ileorectostomised rat model (IRM) is a possible surrogate but evidence as to its validity is scant. In this preliminary study, the resistant starch (RS) content of test breads made from refined low (LAW-R) and high amylose wheat (HAW-R) flours was established in a randomised cross-over trial involving six human ileostomy participants. Starch digestibility of refined breads and diets made from these flours was then evaluated in ileorectostomised rats using a similar experimental format. Physical performance measures and other data were also collected for the rat model. The amount of RS in the low- and high-amylose breads as measured using the human model was 0.8 ± 0.1 and 6.5 ± 0.3 g/100 g, respectively. The RS level of HAW-R bread determined using ileorectostomised rats was 5.5 ± 0.8 g/100 g, about 15% less than that recorded in the human study, whereas for conventional wheat breads the models produced similar RS values. While offering promise, further validation using a wide variety of starchy food products is needed before the IRM can be considered an acceptable alternative for RS determination.

Development and evaluation of chapatti quality of high amylose wheat mutants on the basis of physicochemical, textural and sensory characteristics

Considerable endeavours have been made to develop improved wheat varieties. In fact, starch with increased level of amylose or resistant starch is one of the key research areas that benefits human health. In this study eight high amylose wheat mutant lines including their parent and market flours were assessed for chapatti quality characteristics on the basis of physicochemical (DSC, XRD, RVA), textural, and sensory parameters. Amylose content of mutant lines ranged from ~51 to 75%. Mutant DSC analysis of starch showed variation with increased gelatinization. RVA, XRD, and mixograph data showed variation in starch pasting, crystallinity, and dough characteristics among mutants and parents. 37 other parameters related to grain, starch, flour, dough and chapatti quality were analysed using principal component analysis, ANOVA, and correlation coefficients. Three principal components (PC1, PC2, and PC3) accounted for 68% of the total variation. Strong correlation was observed between amylose and resistant starch content (r2= 0.87). Baking time first side (BTFS) had negative impact on eating quality (EQ) (r2 = −0.81) and pliability was positively correlated to softness (r2 = 0.99). Except for mutant ‘TAC 75’ overall quality scores (49.5–55.5) were as good as or better than parent variety, ‘C 306’ (49.62).

Protein-starch matrix plays a key role in enzymic digestion of high-amylose wheat noodle

Wheat flour, consisting of a complex matrix of starch and protein, is used as a representative model of whole food here to investigate the binary interaction in relation to amylose level and hydrothermal treatment in noodles as a food exemplar. Noodle made of high-amylose wheat (HAW) flour showed an eight-fold higher resistant starch content, compared to the wild type. Protein removal under simulated intestinal digestion conditions resulted in higher starch digestion rate coefficients in raw and cooked flours. In cooked flours, the substrate becomes similarly accessible to digestive enzymes regardless of protein removal. The results indicate that the increased protein content in native HAW flour and thermal stability of starch in HAW noodles lead to higher food integrity and consequently enhance the resistance against α-amylase digestion. Overall, the study suggests that a diversity of starch-protein interactions in wheat-based food products underlies the nutritional value of natural whole foods.

Noodles Made from High Amylose Wheat Flour Attenuate Postprandial Glycaemia in Healthy Adults.

Previous research has not considered the effect of high amylose wheat noodles on postprandial glycaemia. The aim of the study is to investigate the effect of consumption of high amylose noodles on postprandial glycaemia over 2-h periods by monitoring changes in blood glucose concentration and calculating the total area under the blood glucose concentration curve. Twelve healthy young adults were recruited to a repeated measure randomised, single-blinded crossover trial to compare the effect of consuming noodles (180 g) containing 15%, 20% and 45% amylose on postprandial glycaemia. Fasting blood glucose concentrations were taken via finger-prick blood samples. Postprandial blood glucose concentrations were taken at 15, 30, 45, 60, 90 and 120 min. Subjects consuming high amylose noodles made with flour containing 45% amylose had significantly lower blood glucose concentration at 15, 30 and 45 min (5.5 ± 0.11, 6.1 ± 0.11 and 5.6 ± 0.11 mmol/L; p = 0.01) compared to subjects consuming low amylose noodles with 15% amylose (5.8 ± 0.12, 6.6 ± 0.12 and 5.9 ± 0.12 mmol/L). The total area under the blood glucose concentration curve after consumption of high amylose noodles with 45% amylose was 640.4 ± 9.49 mmol/L/min, 3.4% lower than consumption of low amylose noodles with 15% amylose (662.9 ± 9.49 mmol/L/min), p = 0.021. Noodles made from high amylose wheat flour attenuate postprandial glycaemia in healthy young adults, as characterised by the significantly lower blood glucose concentration and a 3.4% reduction in glycaemic response.

Starch granular protein of high-amylose wheat gives innate resistance to amylolysis.

Granular protein is an important structural feature in determining starch digestibility. High-amylose wheat starch (HAWS) with >80% amylose content contains more granular protein than wild-type starch. As analyzed by mass spectrometry-based proteomics, granular-bound starch synthase (GBSS) is the major granular protein in isolated starch materials. GBSS content increases with amylose content (Spearman's correlation, p<0.05), whereas the abundance relative to other proteins is similar among starches. Multiple amylase inhibitors were also identified. From Michaelis-Menten analysis, HAWS has a similar Km (Michaelis constant) as wild type, suggesting initial enzymatic binding is similar. After the pre-digestion of proteins, wild type had a greater change in starch digestibility than HAWS, probably due to the latter having 'thicker' granular-protein layers and higher enzymatic resistance of substrate per se. Overall, the study suggests that the greater granular protein content in HAWS is a factor that contributes to slower amylolysis compared to wild type.

High-amylose wheat starch: Structural basis for water absorption and pasting properties

High-amylose wheat starch (HAWS) and flour (HAWF) have the potential to deliver food products with enhanced nutritional functionality, but structure/function relationships are not well understood. We report the structural bases for differences in water absorption and pasting properties for HAWS and HAWF (amylose contents 71–84 %) compared with wild-type (WTWS/WTWF). With higher amylose content, the proportion of longer amylopectin chains with DP > 25 increased. Both the degree of branching and the branch lengths of amylose were lower for HAWS than WTWS. Compared with WTWF, HAWF contained less total starch, more protein, had lower peak viscosity by high-temperature RVA, lower dough development time and stability time and higher water absorption by Farinograph. Water absorption by HAWS was ∼1.5 times greater than for WTWS, suggesting loose packing of polymers within HAWS granules. Consistent with this, crystallinity and birefringence of starch granules were lower in HAWS.

Complex rheological characterization of normal, waxy and high-amylose wheat lines

• Some lines showed parental characteristics, some behaved completely different. • The samples could be classified into similar groups over the three harvest years. • Some lines were being in the same characteristic group every year, there were stable. • Mixolab and RVA gave similar results. • Correlations were between the amylose/amylopectin ratio and viscous properties.

High-amylose wheat and maize starches have distinctly different granule organization and annealing behaviour: A key role for chain mobility

For regular starches, gelatinization occurs over a relatively narrow temperature range in excess water; however, high-amylose starches typically gelatinise over an extended temperature range by a process that is less understood. The present study compared granule organisation of high-amylose starches from wheat (HAWS) and maize (HAMS), and investigated the role of granule organization in determining the gelatinization heterogeneity and different responses to annealing. During gelatinization (starch/water = 1:2), the melting peaks of double helices and amylose-lipid complexes are merged in HAMS, whereas the peaks are separated for HAWS. It is hypothesized that the glucan chains in HAWS are relatively more mobile (i.e., fewer inter-chain interactions), compared to HAMS. Due to this mobility, hot water leaches amylose more readily from HAWS than from HAMS at similar levels of apparent amylose content. After two-step annealing, To (gelatinization onset temperature) of HAWS increased from 58 to 84 °C, while To of HAMS only increased from 70 to 82 °C. For HAWS, annealing apparently extended double helical lengths and made semi-crystalline lamellae more heterogeneous, but reduced double helix content. For HAMS with more organised granular structure, annealing has a relatively limited effect on lamellar layers and chain conformation. The study demonstrates that HAMS and HAWS have distinctly different granule organisation and behaviours during hydrothermal processing and that HAWS, with more mobile chains, has the potential to form thermostable molecular order. The molecular basis for these differences is proposed to be the greater proportion of very long chains/branches in HAWS than HAMS.

High amylose wheat starch structures display unique fermentability characteristics, microbial community shifts and enzyme degradation profiles.

A slower rate of starch digestion in the small intestine increases the amount of resistant starch (RS) entering the large intestine, which is associated with health benefits. Although increasing the amylose (AM) content of dietary starch intake is one way to increase RS, the processes involved in gut microbial hydrolysis and fermentation of high AM-RS substrates are poorly understood. In this study, five high AM wheat (HAW) starches ranging from 47% AM to 93% AM and a wild type (37% AM), in both native granular and cooked forms, were subjected to in vitro fermentation with a porcine faecal inoculum. Fermentation kinetics, temporal microbial changes, amylolytic enzyme activities and residual starch were determined. All granular starches showed similar fermentation characteristics, independent of AM level, whereas cooking accelerated fermentation of lower AM but slowed fermentation of high AM starches. HAW starches with a very high AM content (>85%) all had similar fermentation kinetics and short-chain fatty acid end-product profiles. Microbial α-amylase, β-amylase, pullulanase and amyloglucosidase enzymatic activities were all detected and followed fermentation kinetics. HAW starch promoted shifts in the microbial community, with increases of the family Lachnospiraceae and the genus Treponema observed, while the genera Prevotella and Streptococcus were reduced in comparison to 37% AM. Overall, these findings suggest that any HAW starch incorporated into high RS food products would be expected to have beneficial microbiota-mediated effects in terms of fermentation kinetics and end products.

Sexually Dimorphic Response of Increasing Dietary Intake of High Amylose Wheat on Metabolic and Reproductive Outcomes in Male and Female Mice

High amylose wheat (HAW) has a higher resistant starch content and lower glycaemic index than standard amylose wheat (SAW), which may be associated with health benefits. This study aimed to determine the effects of replacing SAW with HAW on metabolic and reproductive parameters in male and female mice. Male and female C57BL/6 mice were randomly divided into groups (n = 8/group/sex) and fed either a SAW65 (65% SAW w/w; control), HAW35 (35% HAW w/w), HAW50 (50% HAW w/w) or HAW65 (65% HAW w/w) diet for eight weeks. In male but not female, the HAW65 group had a lower abdominal circumference, relative total fat mass, relative gonadal fat mass and plasma leptin concentration compared to the HAW35 group. There were no differences in fasting blood glucose concentrations or plasma concentrations of cholesterol, triglycerides or non-esterified fatty acids between groups in either males or females. The HAW-fed males had a higher testicular weight and HAW-fed females spent less time in diestrus and a longer time in metestrus compared to the SAW-fed mice. Higher dietary intake of HAW appears to reduce abdominal fat deposition compared to the lower level of HAW in a sexually dimorphic manner. The impacts on reproductive parameters in the HAW-fed mice require further investigation.

High-Amylose Wheat Lowers the Postprandial Glycemic Response to Bread in Healthy Adults: A Randomized Controlled Crossover Trial

Conventional wheat-based foods contain high concentrations of readily digestible starch that commonly give these foods a high postprandial glycemic response and may contribute to the development of type 2 diabetes and cardiovascular disease. The aim of this study was to determine if bread made from high-amylose wheat (HAW) and enriched in resistant starch dampens postprandial glycemia compared with bread made from conventional low-amylose wheat (LAW). This single-center, randomized, double-blinded, crossover controlled study involved 7 consecutive weekly visits. On separate mornings, 20 healthy nondiabetic men and women (mean age 30±3 y; body mass index 23±0.7 kg/m2) consumed a glucose beverage or 4 different breads (each 121 g); LAW-R (refined), LAW-W (wholemeal), HAW-R, or HAW-W. The starch contents of the LAW and HAW breads were 24% and 74% amylose, respectively. Venous blood samples were collected at regular intervals before and for 3 h after the breakfast meal to measure plasma glucose, insulin, ghrelin, and incretin hormone concentrations, and the incremental area under the curve (AUC) was calculated (mmol/L × 3 h). Satiety and cravings were also measured at 30-min intervals during the postprandial period. HAW breads had a glycemic response (AUC) that was 39% less than that achieved with conventional wheat breads (HAW 39±5 mmol/L × 3 h; LAW 64±5 mmol/L × 3 h; P<0.0001). Insulinemic and incretin responses were 24-30% less for HAW breads than for LAW breads (P<0.05). Processing of the flour (wholemeal or refined) did not affect the glycemic, insulinemic, or incretin response. The HAW breads did not influence plasma ghrelin, or subjective measures of satiety or cravings during the postprandial period. Replacing LAW with HAW flour may be an effective strategy for lowering postprandial glycemic and insulinemic responses to bread in healthy men and women, but further research is warranted. This trial was registered at the Australian and New Zealand Clinical Trials Registry as ACTRN12616001289404.

Altering starch branching enzymes in wheat generates high-amylose starch with novel molecular structure and functional properties

Starch that escapes intestinal digestion (resistant starch, RS) is non-glycaemic and considered as dietary fibre. In planta increase of amylose content is a promising approach to make starch resistant to enzyme digestion. Compared to maize, little is known on the effect of amylose content at different length scales of high-amylose wheat (HAW) starch structure and properties. A range of starches with 37%–93% apparent amylose content (AM) were isolated from high-amylose bread wheat produced through non-transgenic breeding altering starch branching enzymes (SBE IIa and IIb). Compared to mutations only in SBEIIa, mutations of both SBE IIa and IIb resulted in higher amylose content with significantly increased proportion of long amylopectin chains (degree of polymerization ≳ 20) and amylose chains. The modified starch molecular structure has significant correlations with starch supramolecular structure and functionality. Compared with wild-type starch, HAW starches showed distinct granular morphology and crystalline pattern with a higher fraction of B-type and V-type polymorph (amylose complex). The peak temperature (Tp) was higher but the onset temperature (To) of gelatinization was not greatly raised (<3 °C) by the mutations. Pasting behaviour of HAW starches (AM<50%) remained similar to wild type, but was greatly reduced at higher AM levels. The data suggest that, depending on food processing condition requirements, different functional and RS properties can be selected based on wheat starch AM content.

Pivotal role of bZIPs in amylose biosynthesis by genome survey and transcriptome analysis in wheat (Triticum aestivum L.) mutants

Starch makes up 70% of the wheat grain, and is an important source of calories for humans, however, the overconsumption of wheat starch may contribute to nutrition-associated health problems. The challenge is to develop resistant starch including high amylose wheat varieties with health benefits. Adapting advance genomic approaches in EMS-induced mutant lines differing in amylose content, basic leucine zipper (bZIP) regulatory factors that may play role in controlling amylose biosynthesis were identified in wheat. bZIP transcription factors are key regulators of starch biosynthesis genes in rice and maize, but their role in regulating these genes in wheat is poorly understood. A genome-wide survey identified 370 wheat bZIPs, clustered in 11 groups, showing variations in amino acids composition and predicted physicochemical properties. Three approaches namely, whole transcriptome sequencing, qRT-PCR, and correlation analysis in contrasting high and low amylose mutants and their parent line identified 24 candidate bZIP (positive and negative regulators), suggesting bZIPs role in high amylose biosynthesis. bZIPs positive role in high amylose biosynthesis is not known. In silico interactome studies of candidate wheat bZIP homologs in Arabidopsis and rice identified their putative functional role. The identified bZIPs are involved in stress-related pathways, flower and seed development, and starch biosynthesis. An in-depth analysis of molecular mechanism of novel candidate bZIPs may help in raising and improving high amylose wheat varieties.

Electrophoretic spectra and activity of peroxidase in winter plants of different varieties

Aim. To study electrophoretic spectra and activity of peroxidase in seedlings of wheat varieties, which differ in genetically determined characteristics. Methods. Biochemicals using a native gel-electrophoresis and a spectrophotometer. Results. Among the studied varieties, the highest peroxidase activity in seedlings varied winter wheat varieties of Astarta, spruce sparrows Polba Golikovskaya and high-amylose wheat HAW. When comparing the molecular forms of the enzyme, it was found that in the seedlings of the studied varieties there were 8–9 isoforms with peroxidase activity, which differed in relative electrophoretic mobility. Inactive isoforms and isoforms with relatively fast mobility were stable and recorded in all studied varieties. The revealed forms with average electrophoretic mobility were in 90 % of the studied varieties. Conclusions. Electrophoretic spectra of peroxidase in seedlings of the studied varieties differ significantly in number and mobility of its multiple molecular forms. Electrophoretic spectra and activity of peroxidase can be used as diagnostic features for comparative analysis of the studied plants in the early phases of ontogeny.&#x0D; Keywords: peroxidase, electrophoretic spectra, varieties, wheat (Triticum L.).

High-Amylose Wheat Foods: A New Opportunity to Meet Dietary Fiber Targets for Health

High-Amylose Wheat Foods: A New Opportunity to Meet Dietary Fiber Targets for Health

High amylose wheat starch increases the resistance to deformation of wheat flour dough

High amylose wheat (HAW) starch has been the focus of a number of nutritional studies, but there is limited information around its effect on the mechanical properties of wheat flour dough. This study investigated the size, shape and packing volume of HAW starch and their effect on the microstructure and rheology of dough. Four flour blends were formulated by adding vital wheat gluten and either HAW or commercial wheat starch to HAW flour to achieve a constant 14% protein content, but varied amounts of HAW starch. A large number of small and irregularly shaped HAW starch granules resulted in a high packing volume per gram of starch. Confocal laser scanning microscopy of optimally mixed doughs correlated the degree of starch granule aggregation with the level of HAW starch in the bi-continuous dough network. Small deformation rheology demonstrated that increased quantities of HAW starch in the dough increased the elastic modulus G′ values. Uniaxial extension measurements highlighted a synergy between HAW starch and sources of gluten proteins resulting in increased strain hardening. The impact of HAW starch on dough rheology was attributed to its irregular shape and large number of small granules leading to greater granule-granule interactions.