Small Starch Granules Research Articles

Functional analysis of ZmMADS1a reveals its role in regulating starch biosynthesis in maize endosperm

MADS-box family proteins play an important role in grain formation and flower development; however, the molecular mechanisms by which transcription factors regulate the starch metabolism pathway are unclear in maize. Here, we report a transcription factor, ZmMADS1a, that controls starch biosynthesis in maize (Zea mays L.). We demonstrate the expression of ZmMADS1a in tassel, silk, and endosperm, and show that the protein is localized to the cell nucleus. Compared with the control, seeds of overexpressing ZmMADS1a increased starch content (especially amylose content), had smaller starch granules and altered chemical structure. Meanwhile, overexpression of ZmMADS1a resulted in increases in the contents of soluble sugars and reducing sugars in maize. ZmMADS1a plays a positive regulatory role in the starch biosynthesis pathway by up-regulating several starch biosynthesis related genes. We also show that ZmMADS1a has a similar adjustment mechanism of starch biosynthesis in rice. Collectively, our study suggests that ZmMADS1a functions as a positive regulator of starch biosynthesis by regulating the expression of key starch metabolism genes during seed development.

Dual-frequency ultrasonic effect on the structure and properties of starch with different size

Single (40 kHz) and dual (40 kHz + 80 kHz) frequency sonication were applied to potato (large granule size) and millet starch (small granule size) at different temperature (25 °C and 60 °C). Starch with large granule size was more sensitive to ultrasound, as reflected by the reduction in peak viscosity, enthalpy, relative crystallinity and degree of order. Ultrasound caused the rough surface, crack and partial collapse on potato starch, however it did not change millet starch significantly. Particle size measurements indicated that the reduction in potato starch particle size was greater than millet starch. The peak viscosity of potato starch decreased by a maximum of 10.7%, whereas the peak viscosity of millet starch decreased by only a maximum of 3.9%. X-ray diffraction showed that the relative crystallinity of potato was reduced by up to 12.3%, however, this change of millet starch was 1.68%. The swelling power of both starches was increased at 25 °C and decreased at 60 °C. Additionally, dual-frequency ultrasound was more effective to modification than single-frequency, irrespective of the granule size of starch. In conclusion, the damage of large granule starch caused by ultrasound was more serious than that of small granule starch and the dual frequency and high temperature will promote the process.

Starch granule size and amylopectin chain length influence starch in vitro enzymatic digestibility in selected rice mutants with similar amylose concentration

In human diet, the products of starch digestion are a major energy source. Starch is stored as water insoluble granules composed of amylose and amylopectin. The susceptibility of starch granule to digestive enzymes is affected by starch granule size, shape, and composition. In this study, starch characteristics and in vitro enzymatic hydrolysis in three rice (Oryza sativa L.) mutants (RSML 184, RSML 278 and RSML 352) with similar amylose concentration (24.3-25.8%) was compared to parent ADT 43 (21.4%). The three mutants had reduced thousand grain weight and starch concentration but higher protein and dietary fiber concentrations. The mutant RSML 352 had small starch granules and reduced short glucan chains [Degree of polymerization (DP) 6-12] compared to the other two mutants (RSML 184 and RSML 278). The mutant RSML 352 had the highest hydrolytic index (HI) and least concentration of resistant starch (RS) compared to the other two mutants and parent rice ADT 43. The two rice mutants (RSML 184 and RSML 278) had reduced HI and increased RS concentration than the parent ADT 43. The results showed that starch granule size and amylopectin structure influence starch enzymatic digestibility and RS concentration.

The effects of field pre-harvest sprouting on the morphological structure and physicochemical properties of rice (Oryza sativa L.) starch.

This study investigates the effects of field pre-harvest sprouting (PHS) on the morphological structure and physicochemical properties of starches from four rice varieties during the rice harvesting period. The results of the study showed that PHS significantly affected the grain characteristics, starch structure, and physicochemical properties of the four rice varieties. Moreover, PHS led to a significant decrease in the grain processing quality and protein content of the four varieties, pitting and unevenness on the surface of the starch granules, and smaller starch granules. In addition, PHS led to starches with lower amylose content, which resulted in a higher swelling power, solubility, and relative crystallinity. Higher gelatinization enthalpy and lower gelatinization temperature were also observed under PHS conditions. The results indicated that PHS can lead to degradation in grain processing and appearance quality, but it can also lead to some improvements in the gelatinization properties of starch.

Inhibition of starch branching enzymes in waxy rice increases the proportion of long branch-chains of amylopectin resulting in the comb-like profiles of starch granules

Starches with comb-like profiles have been detected in some cereal endosperms with inhibiting expression of starch branching enzyme (SBE). Although amylose is considered to be an important factor in the formation of the comb-like profile, the details remain unclear. In this study, a transgenic rice line (GLXN-SBEI/IIb−) was derived from japonica waxy rice cultivar Guang-ling-xiang-nuo (GLXN) through antisense RNA inhibition of both SBEI and SBEIIb. The expression and activity of SBEI, SBEIIb and SBEIIa were declined. The GLXN-SBEI/IIb− endosperm contained large and small starch granules, and these starch granules had the comb-like profiles. The comb-like profiles of starches were detected in GLXN-SBEI/IIb− endosperm after 10 days after flowering with gradually increasing proportion of long branch-chains of amylopectin. The long branch-chains of amylopectin were responsible for forming the comb-like profiles at the outer region of starch granules. The gradually decreasing expression of SBEs influenced the synthesis of amylopectin during endosperm development, resulting in different structure between the inner and outer regions of starch granules from GLXN-SBEI/IIb− endosperm. The above results indicated that the long branch-chains of amylopectin, not amylose, led to the formation of comb-like profiles of starch granules in cereal crops with inhibiting expression of SBEs.

Tarin, a Potential Immunomodulator and COX-Inhibitor Lectin Found in Taro (Colocasia esculenta).

Taro (Colocasia esculenta) corm is a rustic staple food, rich in small starch granules, fibers, and bioactive phytoconstituents such as flavonoids, alkaloids, sterols, tannins, phytates, micronutrients, and proteins, including tarin, a GNA‐related lectin. Tarin exhibits recognized biocide activities against viruses and insects, has antitumoral properties and is an immunomodulator molecule candidate. It has been isolated in highly purified form (>90%) from taro corms through low‐cost and single‐step affinity chromatography. It comprises 2‐domain 27 to 28 kDa protomer, posttranslational cleaved into 2 nonidentical monomers, 11.9 and 12.6 kDa, held by noncovalent binding. At least 10 tarin isoforms sharing over 70% similarity have been described. The monomers assume the β‐prism II fold, consisting of 3 antiparallel β‐sheets formed by 4 β‐strands each. Tarin exhibits an expanded‐binding site for complex and high‐mannose N‐glycan chains 49, 212, 213, 358, 465, and 477 found on cell surface antigens of viruses, insects, cancer, and hematopoietic cells, explaining its broad biological activities. Tarin may stimulate innate and adaptive immune responses, enabling hosts to recover from infections or immunosuppressed status inherent to several pathological conditions. In a murine model, tarin stimulates the in vitro and in vivo proliferation of total spleen and bone marrow cells, especially B lymphocytes. Granulocyte repopulation has also been demonstrated in long‐term mice bone marrow cell cultures. As a potential immunomodulator, tarin, administered to immunosuppressed mice, attenuated cyclophosphamide‐induced leukopenia. We propose a molecular model that unites the potential prophylactic and therapeutic action of tarin on hematopoietic and cancer cells, as a potential immunomodulator.

Extraction and characterization of rye grain starch and its susceptibility to resistant starch formation

The aim of this work was to isolate starch from different parts of rye grain by a traditional and newly developed enzymatic method, and to compare the physical, chemical and molecular properties of isolated starches, especially focusing on susceptibility to resistant starch (RS) formation.Enzymatic treatment provided a high yield of starch which differed in granularity depending on the parts of grain used for isolation. A share of extreme size granules in starches have a strong impact on swelling and pasting properties. The differences could be explained by molecular properties of starch fractions in the examination of which HPSEC with post-column derivatization was especially helpful showing higher dispersity and molar mass of amylose in starches obtained by the enzymatic method.The derivatized SEC profiles of RS have not been reported earlier. Starches isolated by enzymatic method with a high share of small starch granules were less susceptible to a RS formation.

Use of air classification technology to produce protein-enriched barley ingredients

Air classification of an industrial barley endosperm fraction with initial protein content of 8.3% mixed with Aerosil 200F flow aid resulted in protein enrichment up to 28.3% with 21.7% protein separation efficiency. By reducing the classifier wheel speed from 21500 rpm to 8000 rpm, the protein separation efficiency increased to 59.4%, whereas the protein content only decreased to 22.3%. Microstructural analysis showed that small starch granules separated together with protein in the fine fraction and had more continuous protein matrix around them. The protein-enriched fraction obtained at 8000 rpm classifier wheel speed had limited protein solubility of 9.3% at native pH 5.1. Solubility increased in acidic (pH 3, 19.6%) and especially in alkaline (pH 11, 91.9%) conditions. Protein fraction had water and oil binding capacities of 1.2 and 1.1 g/g, respectively. Foaming properties tested at different pH-values revealed best capacity at pH 3. Thermograms showed two melting temperatures, varying from 60.0 to 64.2 °C and from 90.5 to 95.5 °C for all the studied fractions. Cereal side-streams such as the industrial barley endosperm fraction can be utilised as potential plant protein ingredient for various food/feed applications. However, further studies are needed to evaluate the technological, nutritional, and sensory characteristics of this protein enriched fraction.

Small-granule starches from sweet corn and cow cockle: Physical properties and amylopectin branching pattern

Small-granule starches (SGS) attract growing interests with their unique structure and functionalities, such as fat replacement and texture modifications. To explore new resources of SGS, sweet corn starch (SCS) and cow cockle starch (CCS) were studied for their physical properties and branching pattern, with normal corn starch, waxy corn starch, normal rice starch, and waxy rice starch as comparisons. Hydrothermal and wide-angle X-ray powder diffraction analyses showed the much lower content of ordered structure (melting enthalpy of 5.6 J/g) and lower crystallinity (28%) for SCS than for normal corn starch (14.4 J/g and 34%, respectively). The rapid viscosity analyzer analysis showed very low viscosity for SCS through the pasting process and high resistance of CCS against the shear force. The results of β-limit dextrin structure analyses showed highly branched clusters of SCS (about 15 glucan chains per cluster) and a lack of long inter-cluster chains that connect 3 or more clusters. In contrast, CCS displayed a branching pattern similar to that of corn starches, however, with fewer short intra-cluster chains. It was considered that the low crystallinity and viscosity of SCS were attributed to its high branching, which reduced the formation of crystalline structure during the genesis of starch and decreased the interactions among swollen granules. On the other hand, the submicron size made CCS granules less susceptible to the shear force applied during pasting, leading to negligible breakdown. Using SCS and CCS as models, this study connected the physical properties and branching pattern of SGS, thus supporting their potential applications.

Simultaneous silencing of isoamylases ISA1, ISA2 and ISA3 by multi-target RNAi in potato tubers leads to decreased starch content and an early sprouting phenotype.

Isoamylases hydrolyse (1–6)-alpha-D-glucosidic linkages in starch and are involved in both starch granule formation and starch degradation. In plants, three isoamylase isoforms with distinct functions in starch synthesis (ISA1 and ISA2) and degradation (ISA3) have been described. Here, we created transgenic potato plants with simultaneously decreased expression of all three isoamylases using a chimeric RNAi construct targeting all three isoforms. Constitutive expression of the hairpin RNA using the 35S CaMV promoter resulted in efficient silencing of all three isoforms in leaves, growing tubers, and sprouting tubers. Neither plant growth nor tuber yield was effected in isoamylase-deficient potato lines. Interestingly, starch metabolism was found to be impaired in a tissue-specific manner. While leaf starch content was unaffected, tuber starch was significantly reduced. The reduction in tuber starch content in the transgenic plants was accompanied by a decrease in starch granules size, an increased sucrose content and decreased hexose levels. Despite the effects on granule size, only little changes in chain length composition of soluble and insoluble glucose polymers were detected. The transgenic tubers displayed an early sprouting phenotype that was accompanied by an increased level of sucrose in parenchyma cells below the outgrowing bud. Since high sucrose levels promote sprouting, we propose that the increased number of small starch granules may cause an accelerated turnover of glucan chains and hence a more rapid synthesis of sucrose. This observation links alterations in starch structure/degradation with developmental processes like meristem activation and sprout outgrowth in potato tubers.

Both maternally and paternally imprinted genes regulate seed development in rice

Genetic imprinting refers to the unequal expression of paternal and maternal alleles of a gene in sexually reproducing organisms, including mammals and flowering plants. Although many imprinted genes have been identified in plants, the functions of these imprinted genes have remained largely uninvestigated. We report genome-wide analysis of gene expression, DNA methylation and small RNAs in the rice endosperm and functional tests of five imprinted genes during seed development using Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated gene9 (CRISPR/Cas9) gene editing technology. In the rice endosperm, we identified 162 maternally expressed genes (MEGs) and 95 paternally expressed genes (PEGs), which were associated with miniature inverted-repeat transposable elements, imprinted differentially methylated loci and some 21-22 small interfering RNAs (siRNAs) and long noncoding RNAs (lncRNAs). Remarkably, one-third of MEGs and nearly one-half of PEGs were associated with grain yield quantitative trait loci. Most MEGs and some PEGs were expressed specifically in the endosperm. Disruption of two MEGs increased the amount of small starch granules and reduced grain and embryo size, whereas mutation of three PEGs reduced starch content and seed fertility. Our data indicate that both MEGs and PEGs in rice regulate nutrient metabolism and endosperm development, which optimize seed development and offspring fitness to facilitate parental-offspring coadaptation. These imprinted genes and mechanisms could be used to improve the grain yield of rice and other cereal crops.

Effect of Nitrogen Management on the Structure and Physicochemical Properties of Rice Starch.

Nitrogen management (nitrogen application ratio at transplanting, tillering, and panicle initiation growth stages) is an important parameter in crop cultivation and is closely associated with rice yield and grain quality. The physicochemical and structural properties of starches separated from two rice varieties grown under three different nitrogen management ratios (9:1; 7:3; 6:4) were investigated. As the percentage of nitrogen used in the panicle initiation stage increased, the content of small starch granules improved, whereas the content of large granules decreased. Amylose content decreased with increasing nitrogen ratio at the panicle initiation stage, thereby resulting in high swelling power, water solubility, gelatinization enthalpy, and low retrogradation. The X-ray diffraction patterns of the starches were found to be A type. The present study indicated that the best nitrogen management ratio for the cultivation of rice with the highest yield, desirable starch physicochemical properties for high quality cooked rice, and a moderate protein level is 7:3.

Geometrical Formation of Compound Starch Grains in Rice Implements Voronoi Diagram.

Starch forms transparent grains, called starch grains (SGs), in amyloplasts. One of the major morphological SG forms in Poaceae, called a compound SG, is formed by assemblies of small starch granules in an amyloplast. Starch granules assemble as a well-ordered structure; however, the mechanism that regulates this organization has not been identified. In this study, we examined how starch granules grow and converge into the final SG morphology. First, we found that the number of starch granules in an amyloplast is almost constant from the early developmental stage until endosperm maturity. Next, we quantitatively evaluated the geometrical similarities between starch granules and a Voronoi diagram, which is a mathematical tessellation of space based on the distance to a specific set of points in the space. The in silico growth simulation showed that the geometrical patterns of compound SGs resembling a Voronoi diagram is determined by physical interactions among the free-growing starch granules and the amyloplast envelope membrane. The geometrical similarity between compound SGs and a Voronoi diagram is likely a result of maximum loading and storage of starch in the amyloplast. The simulation described in this study provides a greater understanding of how compound SGs are formed and also has the potential to explain morphological variations of SGs.

Effects of Wheat Starch Granule Size Distribution on Qualities of Chinese Steamed Bread and Raw White Noodles

ABSTRACTStarch is a crucial component determining the processing quality of wheat‐based products such as Chinese steamed bread (CSB) and raw white noodles (RWN). Flour from wheat cultivar Zhongmai 175 was used for fractionation into starch, gluten, and water solubles by hand washing. The starch fraction was successfully separated into large (>10 μm diameter) and small starch granules (<10 μm diameter) by repeated sedimentation. Flour fractions were reconstituted to original levels in the flour by using constant gluten and water solubles and varying the weight ratio of large and small starch granules. As the proportion of small granules increased in the reconstituted flours, farinograph water absorption increased, and amylose content, pasting peak viscosity, trough, and final viscosity decreased. Starch granule size distribution significantly affected processing quality of CSB and RWN. Superior crumb structure score (12.0) was observed in CSB made from reconstituted flour with 35% small starch granules. CSB made from reconstituted flours with 30 and 35% small starch granules exhibited the highest total scores, with values of 85.4 and 83.3, respectively. Significant improvements in color, viscoelasticity, and smoothness of RWN were obtained with an increase in small starch granule content, and reconstituted flours with 30–40% small starch granules produced RWN with moderate firmness.

Cytochemical study of developing anthers of Amomum villosum Lour.

We analyzed anther development in Amomum villosum Lour. (Zingiberaceae) using the periodic acid-Schiff's technique and Sudan black staining to test for the presence of starch and lipids, respectively. Our analyses showed that microspore mother cells of A. villosum lack typical callose walls, and numerous lipid granules appear in the cells early in development. Some starch granules are present in anther wall cells, but not in tapetal cells. After meiosis, numerous lipid granules remain unchanged in the microspores. During microspore development, some small starch granules first appear in the central cell region, and then the starch granules increase in size. After microspore division, the bicellular pollen grains become filled with starch and lipids, and remain in this state until the pollen grains reach maturity. At anthesis, the anther wall of A. villosum consists of several layers of endothecium cells with an evidently thickened radial wall, and some layers of parenchyma cells containing numerous starch granules.

Effect of nitrogen fertilizer on distribution of starch granules in different regions of wheat endosperm

This study provided visual evidence of a nitrogen effect on starch granules (SGs) in wheat endosperm. Winter wheat (Titicum aestivum L.) cultivar Xumai 30 was cultured under no nitrogen (control) and 240kgha−1 of nitrogen applied at the booting stage. The number, morphology, and size of A- and B-type SGs in subaleurone of dorsal endosperm (SDE), center of dorsal endosperm (CDE), modified aleurone (MA), subaleurone of ventral endosperm (SVE), and center of ventral endosperm (CVE) were observed under light and electron microscopes. (1) The distribution of SGs in SDE was similar to that in SVE, the distributions of SGs in CDE and CVE were similar, but the distribution of SGs in MA was different from those in the other four endosperm regions. The number of SGs in the five endosperm regions was in the order SDE>CDE>SVE>CVE>MA. (2) Nitrogen increased the number of A- and B-type SGs in SDE and SVE. Nitrogen also increased the number of B-type SGs but decreased the number of A-type SGs in CDE and CVE. Nitrogen decreased the numbers of A-type and B-type SGs in MA. The results suggest that increased N fertilizer application mainly increased the numbers of small SGs and decreased the numbers of large SGs, but that the results varied in different regions of the wheat endosperm.

Mechanical Behavior of Lentil Seeds in Relation to their Physicochemical and Microstructural Characteristics

The mechanical characteristics (rupture force, maximal deformation, and rupture energy) of red and green lentils under compression loading were determined as a function of moisture content ranging from 9.5 to 21.1% (w.b.). Scanning electron microscopy, laser diffraction particle size analysis, and instrumental texture evaluation were successfully applied to relate the microstructure and texture of different lentil seed varieties. Results demonstrated that all of the mechanical parameters of the green lentils, which have smaller starch granules, were significantly (p < 0.05) higher than those of the red lentils. At a loading rate of 4 mm min−1, the force required for initiating seed rupture decreased with an increase in moisture content, for vertical and horizontal orientations (p < 0.05). The scanning electron microscopy observations also revealed that seeds were more flexible in a horizontal orientation.

Starch composition and its granules distribution in wheat grains in relation to post‐anthesis high temperature and drought stress treatments

High temperature (HT) and drought stress (DS) pose serious limitations to wheat (Triticum aestivum L.) grain development, which strongly influences grain yield and quality. In this study, we investigated changes in starch characteristics and starch granules of the high‐yielding winter wheat cultivar Zhengmai 366 in response to HT and DS, both alone and in combination. HT, DS and the combination of these stressors led to a significant reduction in the proportion of A‐type starch granules (&gt;9.9 µm) to B‐type starch granules (&lt;9.9 µm), and the effects of HT and DS were significantly additive. The volume proportion of A‐type starch granules (&gt;9.9 µm) was positively correlated with the contents of AM, AP and starch as well as the grain yield (thousand grains weight) and was negatively correlated with the ratio of AM to AP, whereas there was a contrary correlation between the volume proportion of B‐type starch granules (&lt;9.9 µm) and these values. HT, DS and HT plus DS led to a significant reduction in the content of AP and starch and grain yield, but these treatments had less of an impact on AM content. DS reduced the size of the small starch granules, while HT reduced the size of the large granules. The changes in morphology and size distribution of starch granules led to a reduction in starch content under both stress conditions, consequently reducing grain yield. HT plus DS had a much greater influence on starch characteristics and yield than HT or DS alone.

Structuring and texturing gluten-free pasta: egg albumen or whey proteins?

The effects of adding egg albumen or whey proteins to pasta made from parboiled rice flour (PR) were investigated. Pasta quality was evaluated in terms of color, furosine content, and cooking properties (water absorption, cooking loss, and consistency at the optimal cooking time). The surface heterogeneity of the cooked and uncooked materials was studied, and some starch properties (pasting properties and starch susceptibility to α-amylase hydrolysis) were assessed, along with the features of the protein network as determined by conditional solubility studies and with ultrastructural features of the cooked products. Egg albumen improved pasta appearance and gave a product with low cooking loss, firmer, and nutritionally more valuable than the other ones. In albumen-enriched pasta, small starch granules appear homogeneously surrounded by a protein network. In the uncooked product, the protein network is stabilized mostly by hydrophobic interactions, but additional disulfide interprotein bonds form upon cooking. Thus, addition of 15 % liquid albumen to PR results in significant improvement of the textural and structural features of rice-based gluten-free pasta.

Tracking sulfur and phosphorus within single starch granules using synchrotron X-ray microfluorescence mapping

BackgroundNative starch accumulates as granules containing two glucose polymers: amylose and amylopectin. Phosphate (0.2–0.5%) and proteins (0.1–0.7%) are also present in some starches. Phosphate groups play a major role in starch metabolism while granule-bound starch synthase 1 (GBSS1) which represents up to 95% of the proteins bound to the granule is responsible for amylose biosynthesis. MethodsSynchrotron micro-X-ray fluorescence (μXRF) was used for the first time for high-resolution mapping of GBSS1 and phosphate groups based on the XRF signal of sulfur (S) and phosphorus (P), respectively. Wild-type starches were studied as well as their related mutants lacking GBSS1 or starch-phosphorylating enzyme. ResultsWild-type potato and maize starch exhibited high level of phosphorylation and high content of sulfur respectively when compared to mutant potato starch lacking glucan water dikinase (GWD) and mutant maize starch lacking GBSS1. Phosphate groups are mostly present at the periphery of wild-type potato starch granules, and spread all over the granule in the amylose-free mutant. P and S XRF were also measured within single small starch granules from Arabidopsis or Chlamydomonas not exceeding 3–5μm in diameter. ConclusionsImaging GBSS1 (by S mapping) in potato starch sections showed that the antisense technique suppresses the expression of GBSS1 during biosynthesis. P mapping confirmed that amylose is mostly present in the center of the granule, which had been suggested before. General significanceμXRF is a potentially powerful technique to analyze the minor constituents of starch and understand starch structure/properties or biosynthesis by the use of selected genetic backgrounds.