Starch Chain-length Distributions Research Articles

Morphological, Molecular Structural and Physicochemical Characterization of Starch Granules Formed in Endosperm of Rice with Ectopic Overexpression of α-Amylase.

The objective of this study was to characterize the endosperm starch in rice that ectopically overexpressed the α-amylase. Transgenic rice plants, transformed with cauliflower mosaic virus 35S promoter driven AmyI-1 (35S::AmyI-1) and AmyII-4 (35S::AmyII-4), and 10 kDa prolamin promoter driven AmyI-1 (P10::AmyI-1), were cultivated under standard conditions (23 °C, 12 h in the dark/ 26 °C, 12 h in the light), and brown grains were subsequently harvested. Each grain displayed characteristic chalkiness, while electron microanalyzer (EPMA)-SEM images disclosed numerous small pits on the surface of the starch granules, attributable to α-amylase activity. Fluorescence labeling and capillary electrophoresis analysis of starch chain length distribution revealed no significant alterations in the starches of 35S::AmyI-1 and 35S::AmyII-4 transgenic rice compared to the wild-type. Conversely, the extremely short α-glucan chains (DP 2-8) exhibited a dramatic increase in the P10::AmyI-1 starch. Rapid visco-analyzer analysis also identified variations in the chain length distribution of P10::AmyI-1 starch, manifesting as changes in viscosity. Moreover, 1H-NMR analysis uncovered dynamic modifications in the molecular structure of starch in rice grain transformed with P10::AmyI-1, which was found to possess unprecedented structural characteristics.

Characterization of the starch molecular structure of wheat varying in the content of resistant starch

Resistant starch (RS) is the total amount of starch that is incompletely or not digested and absorbed in the small intestine. It plays a role similar to dietary fibre with beneficial effects for human health. In this study, the RS content of 129 wheat accessions was determined, and the relationship between the several starch physical properties and resistant starch content were analyzed. By comparing the total starch content, amylose starch content, starch chain length distribution, starch crystallization type, starch branching degree, and starch granule morphology between the high RS and low RS content wheat accessions, it was found that the amylose content and RS content were significantly positively correlated. However, in the range of chain length fb 3 (DP ≥ 37), there was a significant negative correlation between amylopectin content and RS content. The surface of starch granules became increasingly smooth as the content of RS increased.

Probing gallic acid–starch interactions through Rapid ViscoAnalyzer in vitro digestion

Phenolic compounds are known inhibitors of starch digestion through binding with α-amylase. However, a growing body of research shows that phenolic-starch interactions at the molecular level may interfere with this inhibition potential. In this study, we evaluated the effect of Gallic Acid (GA) as a model phenolic compound on starch digestion kinetics carried out in vitro in a Rapid ViscoAnalyzer (RVA). The results showed that when GA was added before cooking of starch in order to promote starch-GA complexation, the rate of digestion of starch was similar to that of starch alone, and faster than when GA was added after cooking of starch. The results demonstrated that when GA was introduced after cooking of starch, GA inhibited α-amylase strongly and that inhibition increased with starch paste viscosity only for potato and wheat starches. No correlation was found between starch molecular characteristics and the inhibiting capacity of GA at different starch concentrations. However, the apparent influence of starch chain length distribution suggested that physical effects (such as the absorption of GA at the surface of the starch paste) may play a role in the capacity of GA to inhibit α-amylase.

Starch chain-length distributions determine cooked foxtail millet texture and starch physicochemical properties

Starch chain-length distributions play an important role in controlling cereal product texture and starch physicochemical properties. Cooked foxtail millet texture and starch physicochemical properties were investigated and correlated with starch chain-length distributions in eight foxtail millet varieties. The average chain lengths of amylopectin and amylose were in the range of DP 24–25 and DP 878–1128, respectively. The percentage of short amylopectin chains (Ap1) was negatively correlated with hardness but positively correlated with adhesiveness and cohesion. Conversely, the amount of amylose intermediate chains was positively correlated with hardness but negatively correlated with adhesiveness and cohesion. Additionally, the amount of amylose long chains was negatively correlated with adhesiveness and chewiness. The relative crystallinity (RC) of starch decreased with reductions in the length of amylopectin short chains in foxtail millet. Pasting properties were mainly influenced by the relative length of amylopectin side chains and the percentage of long amylopectin branches (Ap2). Longer amylopectin long chains resulted in lower gelatinization temperature and enthalpy (ΔH). The amount of starch branched chains had important effects on the gelatinization temperature range (ΔT). These results can provide guidance for breeders and food scientists in the selection of foxtail millet with improved quality properties.

A facile method for classifying starch fractions rich in long linear dextrin

Fractionation of linear dextrins with different chain lengths was of great significance for the preparation of functional foods. Traditional gradient non-solvent precipitation method for the fractionation was time-consuming and labor-intensive, making it inconvenient for large-scale dextrin fractionation. The aim of this study was to establish a novel method to fractionate long linear dextrins from debranched waxy maize starch by simply heating (70–80 °C) and centrifuging the debranched starch solution. The results showed that the linear dextrins produced during the debranching process formed ordered structures with melting temperature of 58.8–95.4 °C. According to the analysis of starch thermostability and chain length distribution, it was found that thermostability of ordered structure was significantly and positively correlated with the chain length of linear dextrins. During heating at 70 and 80 °C, less ordered structures with weak thermostability were melted in solution, herein short linear dextrins were removed from debranched starch via centrifugation and thermostable ordered structures could be fractionated to obtain long linear dextrins. By this simple “heating-centrifugation” method, the content of long linear dextrins in starches was increased from 27.08% to 43.40%. This study provided a promising pathway to fractionate long linear dextrins. • A method for isolating long linear dextrin from debranched starch was established. • Ordered structure packed with long linear dextrin (LD) had high thermostability. • Ordered structure was fractionated to obtain long LD by heating and centrifugation. • Long LD content in starch was increased from 27.08% to 43.40% after the treatment. • Short LD with polymerization units of 13–24 formed helical structure rapidly.

Subtle differences in starch fine molecular structure are associated with large differences in texture and digestibility of Chinese steamed bread

Chinese steamed bread (CSB) is a staple food for a large number of people in China, but it has a high glycemic index, which is nutritionally undesirable. Developing a CSB with more slowly digestible starch has been achieved, but current processes for this result in a product with poor sensory appeal. In this study, the relations among starch chain-length distribution, starch digestibility and texture of CSBs made from 10 different wheat flour varieties were investigated, the long-term aim being to develop CSBs with an optimal combination of slow starch digestibility and desirable texture. Although there were only small differences among the CLDs of different wheat flours, they were strongly correlated to the differences of both starch digestibility and textural attributes of CSB. A slower starch digestion rate was associated with relatively longer amylose intermediate to long chains, lower amounts of amylopectin short chains and higher amounts of amylose short chains. A softer CSB texture was related to a relatively higher amount of amylopectin long chains and shorter amylopectin short chains. A higher springiness/resilience of CSB was associated with more amylopectin long chains, longer amylose intermediate to long chains, fewer amylose long chains, and smaller average chain length of amylopectin short chains. A higher cohesiveness of CSB was developed with a higher amount of amylopectin long chains. These results suggest that it is feasible to produce CSB with both slow starch digestibility and desirable texture, by choosing the wheat flour with optimized starch structures. Small variance of wheat starch chain-length distribution can cause a significant difference of starch digestibility and texture of Chinese steamed bread. • Small differences exist among wheat starch chain-length distributions. • Slower starch digestibility associated with higher amounts of amylose short chains. • A softer texture of Chinese steamed bread related to more amylopectin long chains. • Higher springiness of Chinese steamed bread related to fewer amylose long chains.

Indication that starch and sucrose are biomarkers for oil yield in oil palm (Elaeis guineensis Jacq.)

Oil palm (Elaeis guineensis Jacq.) is the most productive oil-producing crop per hectare of land. The oil that accumulates in the mesocarp tissue of the fruit is the highest observed among fruit-producing plants. A comparative analysis between high-, medium-, and low-yielding oil palms, particularly during fruit development, revealed unique characteristics. Metabolomics analysis was able to distinguish accumulation patterns defining of the various developmental stages and oil yield. Interestingly, high- and medium-yielding oil palms exhibited substantially increased sucrose levels compared to low-yielding palms. In addition, parameters such as starch granule morphology, granule size, total starch content, and starch chain length distribution (CLD) differed significantly among the oil yield categories with a clear correlation between oil yield and various starch parameters. These results provide new insights into carbohydrate and starch metabolism for biosynthesis of oil palm fruits, indicating that starch and sucrose can be used as novel, easy-to-analyze, and reliable biomarker for oil yield.

Consecutive reaction kinetics model reveals the nature of long-term rice amylopectin retrogradation characteristics

Starch retrogradation involves nucleation and crystal growth steps, while their relative contribution to the overall retrogradation kinetics and relations with starch fine molecular structures have not been elucidated. In this study, a consecutive reaction kinetics model (CRK) was developed to fit long-term retrogradation kinetics curves for 10 rice starches with distinct molecular structures. Starch chain-length distributions (CLDs) and melting enthalpy kinetics curves for these starches were obtained from our published data. It was shown that these melting enthalpy kinetics curves can be satisfactorily deconvoluted by the CRK model into a combination of nucleation and crystal growth curves. Correlation analysis between CRK model-fitted parameters with starch CLDs showed that starch retrogradation nucleation and crystal growth steps were controlled by distinct starch fine molecular structures. These results have practical applications, as it enables a separate regulation of nucleation and crystal growth steps during retrogradation process of starch-based foods for desirable nutritional properties.

Influence of dynamic high temperature during grain filling on starch fine structure and functional properties of semi-waxy japonica rice

The ripening temperature is a critical factor impacting rice starch structure and properties. In this study, the starch fine structure and functional properties of two semi-waxy japonica rice cultivars were characterized under different dynamic ripening temperatures. Exposure of rice to dynamic high temperature during grain filling increased the relative crystallinity and the lamellar peak intensity of starch. Analysis of starch chain length distribution indicates, as rice grain filling temperature increased, the amylose content (AM) and the proportion of shorter amylopectin chains (DP 6–12 or AP1) decreased, and the proportion of long amylopectin chains (DP 37–76 or AP2) increased, resulting in higher starch gelatinization temperature and enthalpy, as well as pasting viscosities and pasting temperature. The above results imply that high temperature during grain filling can change starch fine structure, and degrade starch gelatinization and pasting properties.

Antagonistic effects of amylopectin and amylose molecules on the starch inter- and intramolecular interactions during retrogradation

Starch retrogradation property is critical for developing functional foods with desirable nutritional and textural attributes, while its nature currently remains elusive. The current study investigated the relations between starch chain-length distributions (CLDs) and interaction patterns (i.e. intermolecular versus intramolecular interactions) of starch molecules during the retrogradation process. CLDs for different waxy potato and sago starch mixtures were characterized by size-exclusion chromatography, and molecular interactions were characterized by differential scanning calorimetry and small amplitude oscillatory shear rheological test. Results for the first time showed that amylose molecules were involved in the formation of short-range intermolecular interactions with amylopectin molecules. Most intermolecularly heterogenous double helices were formed at the 1 : 1 mixing ratio of waxy potato to sago starch. Although the addition of waxy potato starch to sago starch increased the overall amount of short-range double helices, the amount of short-range intermolecular double helices was decreased. It suggests that there was an antagonistic effect between the formation of short-range inter- and intramolecular double helices. These results help a better understanding of the interaction patterns of amylose and amylopectin molecules during the retrogradation process.

In vitro digestion rate of fully gelatinized rice starches is driven by molecular size and amylopectin medium-long chains

In current study, the effects of starch fine molecular structures on its in vitro digestibility at fully gelatinized stage were investigated. The digestion kinetics of 15 fully gelatinized rice starches were obtained and correlated with starch chain-length distributions and molecular size distributions. Both logarithm of slopes and parallel first-order kinetic model were applied to fit the digestion curves to a few kinetics-based parameters. Result showed there were two simultaneous digestion fractions (fast versus slow) for fully gelatinized rice starches. The rate constants of slowly-digestible fraction significantly correlated with starch molecular sizes, especially with that of amylopectin molecules. Hydrodynamically larger amylopectin molecules tend to contain more shorter branches but less long chains. This slows down the starch hydrolysis by α-amylase while the action of AMG is less antagonistically hindered, increasing overall digestion rate. This study provides important information for rice breeders and manufacturers to develop rice products with reduced starch digestibility.

Effects of amylose and amylopectin chain-length distribution on the kinetics of long-term rice starch retrogradation

Understanding the characteristics of long-term starch retrogradation is critical for developing functional foods with desirable textural attributes. The long-term retrogradation kinetics of 10 different rice starches with a wide range of amylose content were thus investigated in this study, focusing on their relations with the starch chain-length distributions (CLDs). Correlation analysis between the retrogradation parameters from differential scanning calorimetry (DSC) with starch CLDs showed for the first time that the amylose content had a parabolic relationship with the melting conclusion temperature and enthalpy of long-term retrograded starches, indicating that the amylose component has a bi-directional effect on the long-term amylopectin retrogradation property. Furthermore, the amount of amylose short-medium chains and amylopectin medium chains were negatively and positively correlated with the melting onset temperature of long-term retrograded starches, respectively. The information learnt from this study could be applied in the rice industry to improve the textural attributes of cooked rice.

Causal relations among starch chain-length distributions, short-term retrogradation and cooked rice texture

Rice is cooked and consumed after a short-term cooling which is associated with the short-term starch retrogradation. The two most important textural attributes (hardness & stickiness) of 28 cooked rice varieties were investigated in this study, based on the causal relations between starch chain-length distributions (CLD) and short-term retrogradation properties. Size-exclusion chromatography (SEC) was used to characterize the starch CLD, while the setback viscosity (SBV) from rapid visco-analyzer and hydrogel firmness (HF) from textural profile analyzer were represented as characteristics of the short-term retrogradation process. Pearson correlations showed that regardless of amylose content, rice starches containing relatively shorter amylose molecules with short to long chains, and more amylopectin medium and long chains with the degree of polymerization (DP) > 69 can all promote the short-term retrogradation rate, resulting in cooked rice with a hard and non-sticky texture. It also suggests that both SBV and HF could be utilized to predict the final cooked rice texture such as the hardness and stickiness. Furthermore, rice starches with a relatively higher amount of medium chains but less short chains within the amylopectin molecules are related to a less sticky texture of the cooked rice. The current study provides important information for rice industry and breeding programs to develop new varieties with targeted eating qualities.

Fecal microbiota responses to rice RS3 are specific to amylose molecular structure

Resistant starch type 3 (RS3) benefits colon health, but the molecular structural reasons for this effect are unclear. Five rice starches with varied amylose content (19.1 %–40.6 %) were used to investigate their effect on gut microbiota. Size-exclusion chromatography and fluorophore-assisted carbohydrate electrophoresis were used to characterize whole starch molecular size distributions and chain length distributions. It was found that RS3 with more chains of degree of polymerization (DP) 36–100 and smaller molecular size can promote the relative abundance of some classes of gut bacteria, while other classes were promoted by RS3 with fewer chains of DP 36–100 and larger molecular size. X-ray diffraction and scanning electron microscopy showed that crystallinity types B or C and differences in physical surface affected the microbiota. This study shows that RS3s with different fine structures are utilized differently by gut microbiota, which may be applied to develop functional foods for gut health.

Insights into chain-length distributions of amylopectin and amylose molecules on the gelatinization property of rice starches

Starch gelatinization is an endothermic transition arising during rice cooking, which significantly influences rice eating and cooking quality (ECQ). The nature of starch (especially amylose) fine molecular structures that gives rise to this endotherm is however currently unclear. A modified Gomperz model was developed in this study to fit the differential scanning calorimetry (DSC) thermograms, resulting complementary information to the traditional DSC parameters. Correlation analysis between DSC parameters with starch chain-length distributions (CLDs) from 14 different rice starches showed for the first time that although amylose CLDs didn't affect starch gelatinization temperatures, the relative length of amylose medium chains was negatively correlated with the gelatinization temperature range. Furthermore, gelatinization onset and peak temperature as well as maximum gelatinization rate were negatively correlated with the relative length of amylopectin short chains, while the conclusion temperature were related to the relative length of amylopectin medium chains. Based on these results, a model for the arrays of amylopectin and amylose molecules within semi-crystalline lamellas of rice starch granules was proposed. These results will enable plant breeders to produce rice with desirable ECQs based on better understandings of the importance of starch fine molecular structures in determining starch gelatinization property.

Effects of Nonstarch Genetic Modifications on Starch Structure and Properties.

This paper examines if, in maize, starch structure and starch-dependent properties might be altered by pleiotropic effects arising from genetic modifications that are not directly related to starch synthesis. The molecular structure, specifically the starch chain-length distributions (CLDs), of two maize lines transformed with Bar (bialaphos resistance) and Cry1c genes (an artificial gene, encoding proteinaceous insecticidal δ-endotoxins) were compared to those of their control lines. The two transgenes are responsible for herbicidal resistance and insect tolerance, respectively. The starch CLDs were measured by enzymatic debranching and measuring the molecular weight distributions of the resulting linear chains. It was found that although all the lines had similar amylose contents, the CLDs of both amylopectin and amylose for Cry1c were noticeably different from the others, having more short amylopectin and long amylose chains. These CLDs are known to affect functional properties, and indeed it was found that the Cry1c transgenic lines showed a lower gelatinization temperature and faster digestion rate than the control or Bar lines. However, a slower digestion rate is nutritionally desirable. Thus, pleiotropic effects from genetic modifications can indirectly but significantly affect the starch synthesis pathway and thus change functional properties of significance for human health.

Distribution of short to medium amylose chains are major controllers of in vitro digestion of retrograded rice starch

In many cultures, rice is let to cool for a while after cooking before being consumed, which results in some retrogradation of the starch. The in vitro digestibility of 16 cooked rice starches after extended cold (4 °C) storage, which leads to extensive retrogradation, was investigated from the perspective of their starch molecular fine structure. Size-exclusion chromatography (SEC) and fluorophore-assisted carbohydrate electrophoresis (FACE) were used to characterize the starch chain-length distributions (CLDs) and whole molecular size distributions. The retrograded starch gel network was studied by differential scanning calorimetry and scanning electron microscopy. Pearson correlation analysis with in vitro digestibility revealed that the starch digestion rate was positively correlated with cell size and cell wall thickness within the formed gel network, which was mainly derived from the interactions of amylose short-medium chains and not strongly related to amylopectin molecular structure. This suggests that rather than just altering amylose content, modification of amylose fine molecular structure (e.g. increasing the amount of short-medium amylose chains) can also slow rice starch digestibility.

A mutation in Waxy gene affects amylose content, starch granules and kernel characteristics of barley (Hordeum vulgare)

AbstractWaxy barley referred to as low‐amylose or amylose‐free has special advantages in nutrition composition and food processing. Waxy gene encoding granule–bound starch synthase I (GBSSI) is responsible for amylose synthesis in barley. The G3935‐to‐T in Waxy gene has been previously found in amylose–free barley. In this study, G3935‐to‐T was proved to co‐segregate with the waxy phenotype of barley, but has no obvious effect on GBSSI catalytic activity and starch chain length distribution. However, recombinant inbred lines with G3935‐to‐T in Waxy gene are of significant modification in starch granules morphology and pasting properties, increase of grain β‐glucan content, and decrease of thousand kernel weight along with lower kernel width. A polymerase chain reaction with confronting two–pair primers marker was developed for economic and efficient screening of G3935‐to‐T. This study provides the basis for cultivar improvement of waxy barley then fully developing its potential value and utility in food processing.

The multi-scale structure, thermal and digestion properties of mung bean starch.

Identification and selection one special variety mung bean for lower GI food is very useful, however, the fundamental study for mung bean starch is still insufficient to meet its demand. In this study, four varieties of mostly planted mung bean in China were selected as model materials. The multi-scale structure of mung bean starch was characterized by SEC, HPAEC, XRD, SAXS, and SEM. SEC and HPAEC give the amylose contents, amylose and amylopectin fine structure of mung bean starch. Mung bean starch from XRD spectrum display CA type semi crystallinity. The crystalline lamellar thickness from SAXS curves were 7.34-7.60 nm. DSC indicated that the peak gelatinization temperature is at 67 °C-68 °C. Resistant starch in mung bean disappears rapidly after cooking, although the amount of slowly digested starch was still more than half of the total starch. Since the gene backgrounds of the mung bean starch samples are very close, there was no obvious difference in their molecular and aggregated state structure, and the digestion properties were similar, too. Unique SEC and HPAEC profiles of starch chain length distribution can be utilized to help find more genetic resources and cultivate variety to meet the needs for starch applications.

Editing of Rice Isoamylase Gene ISA1 Provides Insights into Its Function in Starch Formation

Isoamylase 1 (ISA1) is an isoamylase-type debranching enzyme which plays a predominant role in amylopectin synthesis. In this study, the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated endonuclease 9 (CRISPR/Cas9) system was used to edit ISA1 gene in rice via Agrobacterium-mediated transformation. We identified 36 genetic edited lines from 55 T0 transgenic events, and classified the mutation forms into 7 types. Of those, two homozygous mutants, cr-isa1-1 (type 1, with an adenine insertion) and cr-isa1-2 (type 3, with a cytosine deletion) were selected for further analysis. Seed sizes of both cr-isa1-1 and cr-isa1-2 were affected, and the two mutants also displayed a shrunken endosperm with significantly lower grain weight. Electron microscopy analysis showed that abnormal starch granules and amyloplasts were found in cr-isa1-1 and cr-isa1-2 endosperm cells. The contents of total starch, amylose and amylopectin in the endosperm of the cr-isa1 mutants were significantly reduced, whereas sugar content and starch gel consistency were observably increased compared to the wild-type. The gelatinization temperature and starch chain length distributions of the cr-isa1 mutants were also altered. Moreover, transcript levels of most starch synthesis-related genes were significantly lower in cr-isa1 mutants. In conclusion, the results indicated that gene edition of ISA1 affected starch synthesis and endosperm development, and brought potential implications for rice quality breeding.