Comparison of the GFFF and LALLS Methods for the Measurement of Starch Granule Size Distribution in Spring Barley Caryopses

A newly developed method GFFF (Gravitational Field‐Flow Fractionation) and the well known method LALLS (Low Angle Laser Light Scattering) were used to assess starch granule size distribution of ten varieties of spring barley. As a distribution criterion, the ratio of starch granule content larger than 8 μm (type A) and smaller than 8 μm (type B) was chosen. Both methods divided the observed set in a similar way. Varieties Akcent, Forum and Atribut formed a variety set with the highest ratio of large and small starch granules. Varieties Scarlet and Kompakt had intermediate ratios. The remaining five varieties Amulet, Novum, Olbram, Tolar and Krona had the lowest ratios of large and small starch granules. Statistical analysis showed that there was a highly significant correlation between the GFFF and LALLS methods.     

Monitoring of barley starch amylolysis by gravitational field flow fractionation and MALDI-TOF MS

BACKGROUND: In barley, starch occurs in the form of granules with bimodal size distribution. Enzymatic hydrolysis of the starch granule is one of the most important reactions occurring during malting and mashing. Previous studies revealed the discrepancies in the assumption that barley varieties with better malting qualities should have a higher A/B (large/small starch granules) ratio. This led us to focus our attention on detailed analysis of two barley varieties, Jersey and Tolar, both with high malting quality but significantly differing in A/B (1.28 and 0.66, respectively), were chosen for more detailed analysis in the actual work. In this study, the capacity of gravitational field flow fractionation (GFFF) to monitor amylolysis of the starch granules was investigated. RESULTS: Isolated starch granules from these two barley cultivars were treated with amylases. The changes in starch granule size and bimodal distribution were studied by GFFF. Simultaneously, free sugars released during enzymatic digestion were observed by matrix‐assisted laser desorption ionization time‐of‐flight mass spectrometry. The changes in the fractogram and in the mass spectra reflect a correlation with the progress of enzymatic hydrolysis. CONCLUSION: The results show the interest in utilization of GFFF as a simple and cheap method for monitoring changes in the distribution of the starch granule size during amylolysis. Copyright © 2011 Society of Chemical Industry     

Differential Binding of Concanavilin A to Starch Granules Isolated from Barley and Maize

The location of amylose and amylopectin components and/or the degree of branching at the exterior of starch granules may influence the binding of F. I. T. C. labelled Concanavilin A. Waxy Maize starch granules and small starch granules from barley have a higher affinity for lectin than starch granules isolated from normal maize or large starch granules isolated from barley kernels. The binding of Concanavilin A to starch granules is shown to be cooperative and reversible.     

Microstructure of Buckwheat and Barley During Malting Observed by Confocal Scanning Laser Microscopy and Scanning Electron Microscopy

The structures of buckwheat and barley were observed by Scanning Electron Microscopy (SEM) and Confocal Scanning Laser Microscopy (CSLM) throughout the malting process. SEM and CSLM have different merits when studying grains and seeds. SEM can be used to study the detailed structure (high magnifications can be achieved) and CSLM to study the overall change in structure during processing. SEM proved that buckwheat starch is degraded by both pitting and surface erosion. A concentric sphere structure was visible when buckwheat starch granules were partly broken down. With CSLM, the organization of protein and starch of buckwheat and barley could be examined independently in projections and images visualizing the 3D structure. CSLM proved that buckwheat starch is surrounded by a protein matrix. Buckwheat protein was structured and surrounded starch granules, while barley protein was present in an amorphous mass through the endosperm. The differences in overall structure between unmalted and malted seeds were evident, as the starch and protein in both the barley and buckwheat were degraded. CSLM may be used in the future to couple processing characteristics to the structure of grains and seeds.     

α-AMYLASE IN DEVELOPING BARLEY KERNELS — A REAPPRAISAL1

The outer layers that can be dissected readily from immature barley kernels were identified by light microscopy. α-Amylase was located in the outer pericarp of developing Bonanza and Himalaya barley. Only low pl α-amylase (α-amylase 1) was detected in the pericarps but only one of the two major low pl components of α-amylase from germinated Himalaya was found in the pericarp tissue. These enzymes appeared to hydrolyze the small starch granules (2–3 μm) present in the outer pericarp of developing barley kernels. The even smaller starch granules (0.25–2μm) present in the inner pericarp were hydrolyzed at a later stage of kernel development.     

A Reassessment of the Chemical Structure of Barley and Wheat Starch Granules

Barley starch exists in two clearly defined populations of large (ca. 25 μ diameter) and small (ca. 5, μ diameter) granules, whilst in wheat there is a range of granule sizes with no similar bimodal distribution. The small granules of barley represent about 90% of the total starch granules by number, but account for only about 10% of the total starch in weight. Fractions containing only large or only small granules have been isolated from both barley and wheat. The small granules from barley have a higher amylose content than the large granules, a higher gelatinisation temperature, and are usually associated with more protein. In contrast, large and small granules from wheat differ only slightly in their respective amylose contents but, like barley differ in gelatinisation temperatures and amounts of associated protein. It is proposed that starch synthesis in the small starch granules of barley is under different genetic control from that in the large granules.     

SITE OF α-AMYLASE IN DEVELOPING BARLEY KERNELS

Dissection studies of two cultivars of barley showed that the pericarp of developing barley kernels contained α-amylase as well as starch granules. Shortly after anthesis, the starch granules disappeared completely and the α-amylase activity of the pericarp fell rapidly to a low level that was maintained until the kernels reached maturity. The α-amylase was isolated, partially purified by ion-exchange chromatography on carboxymethyl (CM) cellulose followed by selective precipitation with glycogen, and characterized by analysing the end-products of its action on amylose.     

Variations in morphology and composition of barley endosperm cell walls

BACKGROUND: The endosperm cell walls (CWs) impact upon barley grain utilization as well as influence animal and human nutrition. The objective of this study was to examine the morphological and compositional differences in the endosperm CWs derived from barley grain grown in different (uncontrolled) environments in Canada, and differing in grain hardness, protein, and total β‐glucan contents. RESULTS: The endosperm CWs were isolated from barley, cv. Metcalfe, grown in Davidson, SK (sample A), Hythe, AB (sample B), and Hamiota, MB (sample C). The inner wall surfaces of sample A exhibited deep indentations made by large and small starch granules, whereas the CWs of sample B were thicker, had smooth surface, and exhibited a greater degree of binding between the adjoining cells. The CWs of sample A contained the lowest amount of β‐glucans but the highest amount of arabinoxylans and the mannose‐containing (glucomannans) polysaccharides. The CWs of sample B contained the highest ratio of ferulic and dehydrodiferulate residues to Xyl residues among the samples, indicating the greatest potential and degree of cross‐linking of arabinoxylans in these preparations which might be related to the strength and rigidity of the grain endosperm. The amino acid composition of the proteins detected in the isolated CW indicated the presence of 37–47 g kg^?1 of structural proteins. CONCLUSIONS: Substantial differences in the morphological appearance and chemical composition of the CW preparations were observed and some attempts were made to relate these differences to the variable composition and physical properties of barley grain as affected by the environmental factors. Copyright © 2008 Crown in the Right of Canada, Canadian Grain Commission. Published by John Wiley and Sons, Ltd     

A CRITICAL EXAMINATION OF PROCEDURES FOR THE ISOLATION OF BARLEY STARCH

A study has been made of the methods available for isolating and purifying barley starch granules. A detailed examination involving light and scanning electron microscopy and Coulter counter analysis has indicated that discarding the brown protein layer, which appears on top of centrifuged suspensions of crude starch, results in severe loss of small granules. A protocol is suggested whereby this brown layer is purified separately and then added back to the white layer to give representative starch granule preparations without loss of small granules.     

A Study of Starch Granule Size and Distribution in 29 Barley Varieties

The starch granule size and distribution has been determined on 29 different samples of barley. There is a wide range in the ratio of small to large granules which varies from a minimum of 5.5:1 to a maximum of 37:1. Small granules were separated from large granules in four varieties and the average weight of the granules determined. From these values the actual percentage of small granules in the original barley starch was determined. Small granule starch may account for from 6.2–30.6% of total starch weight in the varieties examined.     

In situ Raman microscopy of starch granule structures in wild type and ae mutant maize kernels

A method developed for in situ imaging of starch granule structure in dry seeds has been applied to compare the starch granule structures found in wild type and ae mutant maize kernels. In the isogenic ae mutant the activity of the starch branching enzyme IIb is inhibited, which gives rise to a high amylose starch. The granule structures in the wild type samples have been found to be homogeneous, whereas those in the ae mutant are grossly heterogeneous within individual granules, between granules within individual cells, and between cells across the endosperm. The level of heterogeneity observed in situ appears to be more marked than that previously reported for studies on isolated ae mutant starches. Iodine/potassium iodide staining and polarised light microscopy have been used together with Raman microscopy, which has allowed high‐resolution mapping of the composition and physical state of the structures within the granules, to probe the origins of the heterogeneity of the starch structures. Although the mutation inhibits the activity of the branching enzyme within the granules, and both the composition and level of crystallinity within and between granules is variable, the major origin of the heterogeneity of the granule architecture appears to result from significant changes in the assembly and packaging of the crystalline structures within the granule. It is suggested that this arises due to the mutation of the starch branching enzyme introducing defects into the self‐assembly of the crystalline structure, resulting in an accumulation of defects and increased randomisation of the granule structure.     

高直链玉米III型抗性淀粉制备及其结构和特性

以高直链玉米淀粉G50和G70为原料,经酸解、糊化、脱支和重结晶步骤获得III型抗性淀粉,通过退火与压热处理以进一步提升淀粉的抗性比例。采用扫描电子显微镜、X射线衍射、差示扫描量热、快速黏度分析等方法,研究淀粉颗粒形貌、结晶结构、热特性及糊化特性,利用Englyst法测试淀粉消化特性。结果表明：高直链玉米淀粉G50和G70酸解后的得率分别为77.9%和84.5%,重结晶后的得率降为54.4%和70.2%。原G50和G70改性后,淀粉颗粒形貌被破坏,形成大小不等、颗粒形貌不规则的团聚体;淀粉结晶型由B＋V型转变为A＋V型,且结晶度升高;淀粉糊化温度升高,且加热过程中黏度几乎消失。溶解与膨胀特性结果表明,经酸解、糊化、脱支和老化处理后原G50和G70的溶解性显著升高,退火和压热处理后降低了III型抗性淀粉的溶解性和膨胀度。体外消化特性分析表明,改性后的G50和G70具备更强的抗消化性能,抗性淀粉含量最高可达80.5%（G70-RS3-压热20%）。本研究的改性处理能有效提高高直链玉米淀粉G50和G70中抗性淀粉含量,同时抗性淀粉含量与结晶度和糊化温度呈显著正相关。     

茶多酚-高直链玉米淀粉共研磨混合物的制备与结构表征

为了提高抗性淀粉的含量,以茶多酚(TPs)和高直链玉米淀粉(HAMS)为原料,利用研磨技术制备茶多酚-淀粉共研磨混合物,通过X射线衍射、扫描电子显微镜和热重分析对其结构、形貌和热稳定性进行表征,并采用模拟体外消化方法对其营养片段(快消化淀粉、慢消化淀粉、抗性淀粉)进行评价。X-射线衍射表明,随着球磨时间的延长,所得共研物中淀粉的结晶衍射峰强度逐渐减弱,经3 h球磨的处理后,淀粉的结晶度由38.1%降低到8.3%。扫描电镜结果表明:淀粉颗粒膨胀,颗粒形状发生形变,表面变得相对粗糙,部分颗粒发生破裂,茶多酚与淀粉在共研磨过程中发生聚集粘连。热重分析结果表明,所得共研磨物较高直链玉米淀粉的稳定性低,但比茶多酚的热稳定性高。经共研磨处理后,当TPs与HAMS质量比由1:50增加到1:10时,所得共研磨物的抗性淀粉含量由6.31%±0.88%增加到31.92%±1.53%。经过共研磨处理后茶多酚-高直链玉米淀粉共研磨混合物的结晶度降低,形态发生变化,表面变得粗糙,颗粒黏连,热稳定性降低,抗性淀粉的含量增加。     

Scanning electron microscopy of Fusarium damaged kernels of spring wheat

Kernels of five wheat cultivars (Triticum aestivum) of different bread-making quality were examined. Grown under field conditions, heads of wheat were inoculated in the flowering stage with an aqueous suspension of Fusarium culmorum conidia. Wheat heads were collected from the control and inoculated plots at full maturity. Control (non-inoculated) kernels without any symptoms of disease and Fusarium damaged kernels (FDK) were examined under scanning electron microscopy (SEM). Examination of the FDK fraction confirmed localisation of Fusarium hyphae on the surface and inside the tissues of kernels. Observations of the endosperm from Fusarium infected kernels revealed presence of fungal hyphae in the endosperm and some characteristic structural changes in many of its regions, such as partial or complete lack of the protein matrix, damage to large and small starch granules caused by fungal amylolytic enzymes, disappearance of small starch granules as the colonisation progressed, complete disappearance of the starchy endosperm under severe infection. Fungal colonisation of the endosperm and structural changes in its area were highly variable traits within the FDK fraction of a given cultivar.     

The size distribution of starch granules in endosperm of different sized kernels of the wheat cultivar maris huntsman

The particle size distributions of starches prepared by proteolysis from different sized kernels of Maris Huntsman wheat, were determined by Coulter Counter. The number of starch granules was greater in “large plump” kernels than in “small plump” and “shrivelled” kernels grown under the same conditions. In all three kernel types more than one third of the total starch weight was contributed by granules less than 10 μm diameter (B type granules). The proportion of these small granules was significantly greater in “large plump” kernels than in the other two types.     

The Effect of Physical Damage on Large and Small Barley Starch Granules

Barley starch from a Scottish brewing barley (Golden Promise) has been isolated, purified and fractionated into two populations according to the granule sizes. The effects of physical damage on the two types of granules have been examined by iodine staining, debranching, gel chromatography, viscosity measurements and scanning electron microscopy. The amylose content of large starch granules was slightly greater than that of the small granules and the effect of physical damage did not differ significantly between the two types of granules. Physical damage to both types of granules in the dry state followed by extraction with cold water resulted in the preferential solubilization of low molecular weight amylopectin.     

小麦A、B型淀粉分离提取方法研究

该研究对小麦A、B型淀粉分离提取方法进行探讨,结果表明,采用水柱沉淀离心法,在淀粉浆浓度0.1 g/mL、离心速度200 r/min、静置时间40 min和静置次数3次条件下,可快速、有效分离A、B型淀粉;A、B型淀粉得率分别为74.2%和23.7%,淀粉损失2.1%。经显微照相显示,此条件下A、B型淀粉分离彻底。     

四种元麦淀粉粒形态结构与理化性质的比较

为探明不同品种元麦淀粉粒形态结构与理化性质的差异,以通麦6号、苏裸麦2号、青元麦、黑元麦为实验材料,采用扫描电镜观察、X-射线衍射、傅里叶变换远红外光谱等方法研究了这四类元麦淀粉粒形态特征及其理化性质,结果表明:青元麦,苏裸麦2号和通麦6号淀粉颗粒比较光滑,大小比较均匀,大颗粒多为圆饼形,小颗粒多为圆球形。黑元麦颗粒大小不均匀,差异较大。4类元麦的表观直链淀粉含量不同,其中黑元麦的直链淀粉含量显著高于其他品种;黑元麦和苏裸麦2号的B-型淀粉粒较多,通麦6号和青元麦的A-型淀粉粒较多。黑元麦的淀粉相对结晶度和表层结构有序度最高,溶解度最高、膨胀势最低。在淀粉葡萄糖苷酶水解过程中,青元麦水解程度最高,苏裸麦2号水解程度最低;在猪胰腺α-淀粉酶水解过程中,通麦6号水解程度最高,青元麦最低;在HCl水解过程中,青元麦水解程度最高,黑元麦水解程度最低。     

Effects of Granule Sizes on Physicochemical Properties of Cross‐linked and Acetylated Wheat Starches

Large and small wheat starch granules were used for cross‐linking and acetylation to determine effects of granule sizes on physicochemical properties of the modified starches. The native and cross‐linked starches from the small granules showed higher phosphorus contents than did those from the large granules. However, the level of phosphate substituents in the modified starches was not significantly different between the large and small granules under the same conditions. In contrast, the large granules had a higher reactivity with acetic anhydride than did the small granules. The phosphate group cross‐linked starch (CS), acetylated starch (AS) and acetylated cross‐linked starch (ACS) from the large granules had lower gelatinization temperatures and higher enthalpies than those from the small granules. The paste viscosities of the CSs from the large granules decreased rapidly, whereas those of the AS or ACS increased significantly as compared with those from the small granules. The pastes of cross‐linked starches from the small granules were more stable than those from the large granules, whereas the pastes of AS and ACS from the large and small granules had similar resistance to freeze‐thaw treatment. Scanning electron microscopy (SEM) also showed that the small granules were less damaged after modification than the large ones. Thus, the different granule sizes resulted in different physicochemical properties of starch after modification.     

THE IN VIVO AND IN VITRO DEGRADATION OF BARLEY AND MALT STARCH GRANULES

The susceptibilities to amylolytic hydrolysis of the two different types of starch granule in barley and malt have been investigated. The large and small granules from both germinated and ungerminated grain were subjected to the sole action of malt α-amylase under conditions which otherwise simulated those of a conventional infusion mash. Large starch granules from barley are hydrolysed at a slower rate than those from malt. The faster conversion of the latter is attributed to prior modification of the starch granule structure during germination. The small granules from barley are extremely resistant to attack by α-amylase and even pre-cooking does not appreciably increase their susceptibility to amylase attack. Although the small starch granules from malt are less susceptible than the corresponding large granule fraction, they can be hydrolysed to a greater extent than can the small granules from barley. The increased susceptibility of small granules from malt is probably due to the partial removal of their protecting surface layer of protein. Although during malting the small granules of barley are hydrolysed more rapidly than the large granules, the situation is reversed during mashing. Very little loss of extract can be attributed to the enzymic resistance of small starch granules in all-malt mashes. If raw barley is used in the grist, substantial amounts of small starch granules may remain in the mash.