Fine Structure Of Amylopectin Research Articles

The effects of chain-length distributions on starch-related properties in waxy rices

Normal rice starch consists of amylopectin and amylose, whose relative amounts and chain-length distributions (CLDs) are major determinants of the digestibility and rheology of cooked rice, and are related to metabolic health and consumer preference. Here, the mechanism of how molecular structural features of pure amylopectin (waxy) starches affect starch properties was explored. Following debranching, chain-length distributions of seven waxy varieties were measured using size-exclusion chromatography, and parameterized using biosynthesis-based models, which involve breaking up the chain-length distribution into contributions from five enzyme sets covering overlapping ranges of chain length; structure-property correlations involving the fifth set were found to be statistically significant. Digestibility was measured in vitro, and parameters for the slower and longer digestion phase quantified using non-linear least-squares fitting. The coefficient for the significant correlation involving amylopectin fine structure for the fifth set was −0.903, while the amounts of amylopectin short and long chains were found to dominate breakdown viscosity (correlation coefficients 0.801 and − 0.911, respectively). This provides a methodology for finding or developing healthier starch in terms of lower digestion rate, while also having acceptable palatability. As rice breeders can to some extent control CLDs, this can help the development of waxy rices with improved properties.

The relationship between starch synthesis enzyme activity, gene expression, and amylopectin fine structure in waxy maize

The relationship between starch synthesis enzyme activity, gene expression, and amylopectin fine structure in waxy maize

Comparative analysis of granular starch hydrolysis and multi-structural changes by diverse α-amylases sources: Insights from waxy rice starch

Three cultivars of waxy rice starch with different multi-scale structures were subjected to α-amylase hydrolysis to determine amylopectin fine structure, production of oligosaccharides, morphology, and crystallinity of the partially hydrolyzed starch granules. α-amylases hydrolyzed the amylopectin B2 chain during the initial stage of hydrolysis, suggesting that it is primarily located in the outer shell of the granules. For waxy rice starch with loose structure, α-amylases attacked the crystalline and amorphous regions simultaneously in the initial stage, while for starch granules with compact structure, the outer shell blocklet (crystalline structure) can be a hurdle for α-amylases to proceed to hydrolysis of the internal granule structure. The ability of α-amylases from porcine pancreatic α-amylases to attack the outer shell crystalline structure was lower than that of α-amylases from Bacillus amyloliquefaciens and Aspergillus oryzae. These results show that α-amylase source and rice cultivar combinations can be used to generate diverse structures in degraded waxy rice starch.

The relationship between the fine structure of amylopectin and the type of crystalline allomorph of starch granules in rice endosperm

AbstractBackground and ObjectivesIt is known that any wild‐type varieties of cereals so far examined have A‐type starch crystals in their endosperm whereas their starch branching enzyme 2b (be2b) mutants that are usually referred to as amylose‐extender (ae) mutants produce B‐type starch crystals. The present study aimed to examine the structural features of amylopectin which are responsible for the A‐type or B‐type crystalline allomorphs of starch granules by using starches in wild‐type japonica and indica rice varieties and their be2b (ae) mutants.FindingsThe average length of chains of A‐type amylopectin was markedly shorter than that of B‐type amylopectin. It was also thought that A‐type amylopectin had two types of branches, namely the first branches were formed mainly by BEI in the basal region of the cluster and the second branches were synthesized specifically by BEIIb inside the cluster. These differences caused the average length of double helices formed by cluster constituent chains to be shorter in wild‐types than that in their be2b mutants, and these differences changed the crystalline allomorph of starch granules to B‐type in the mutants from A‐type in their wild‐types, as examined by wide angle X‐ray diffraction and sum frequency generation spectroscopy.ConclusionsCombined with these observations and calculations of chain‐length of hypothetical A chains in four types of amylopectin molecules, it was concluded that both the average length of external segments of cluster chains and the formation of the second branches inside the cluster greatly affect the crystal types of starch granules.Significance and NoveltyThe novel findings in the present study can provide new insights into the structural features of amylopectin which determine the A‐type or B‐type crystalline allomorph of starch granules of cereals.

Generating waxy rice starch with target type of amylopectin fine structure and gelatinization temperature by waxy gene editing

Waxy rice, which lacks amylose, is an important variant in rice, and its starches have been widely used. New waxy rice varieties generated via the CRISPR/Cas9 gene-editing system is described. Herein, four waxy rice starches with different physicochemical properties were successfully obtained by the CRISPR/Cas9 editing Waxy (Wx) gene. The results showed that the amylose content (AC) of wx mutant starches ranged from 0.26 to 1.78 %, and CZBwx1 starches had the best gel consistency and highest water solubility among all wx mutants. Mutations of Wxb allele produced more short-chains (degree of chain polymerization (DP) 6–11), and less medium- and long-chains (DP12–70) than that of Wxa, while the AC of Wxa allele mutants was higher than the mutations of Wxb allele. The gelatinization temperature (GT) of wxa mutant starches was higher than that of wxb mutant starches. Moreover, we found that the GT and amylopectin fine structure type of waxy rice starch did not change after Wx gene editing. Based on these findings, it is possible to produce waxy rice starch with different physicochemical properties, containing target GT and chain length distributions of amylopectin.

Evaluation of in vitro digestion methods and starch structure components as determinants for predicting the glycemic index of rice

Mainstreaming the low glycemic index (GI) trait in breeding programs is constrained by low-throughput and high-cost clinical GI phenotyping. This study aimed to evaluate the potential of starch fine structure components and simulated digestion parameters in predicting GI in rice. Amylose (AM1 and AM2; r = −0.94 and r = −0.80, respectively, p < .05) and amylopectin fine structure (MCAP, SCAP, and SCAP1; r = 0.78-0.86, p < .05) measured through size-exclusion chromatography along with resistant starch (r = −0.81, p < .05) in seven (7) rice accessions showed high correlation with in vivo GI. Meanwhile, starch hydrolysis extent (SH) and the corresponding area under the digestion curve (AUC) obtained through in vitro digestion were found to be of higher correlation with GI, even within shorter digestion periods of 5 min or 30 min (r = 0.96, p < .01). These results highlight the potential use of these parameters as predictors of GI, with improved predictive capacity through a multiple regression model. Higher correlations of simulated digestion AUC with GI may be due to its ability to account for the overall food matrix native macro- and micro-structures, gaining an added advantage over SEC method as a predictive tool in studying rice GI variability. Validation in a larger population is an inevitable next step.

Assessment of the Characteristics of Waxy Rice Mutants Generated by CRISPR/Cas9.

The cooking and eating quality of rice grains is a major focus from a consumer’s perspective and is mainly determined by the apparent amylose content (AAC) of the starch. Waxy rice, a type of rice with an AAC of less than 2%, is an important goal for the breeding of high-quality rice. In recent years, the cloning of the Waxy (Wx) gene has revealed the molecular mechanism of the formation of waxy traits in rice. However, there have been limited studies on the physicochemical properties, such as gelatinization temperature, rapid viscosity analyzer profile, and amylopectin fine structure of wx mutants. In the current study, a rapid and highly efficient strategy was developed through the CRISPR/Cas9 gene-editing system for generating wx mutants in the background of five different rice varieties. The wx mutation significantly reduced the AAC and starch viscosity but did not affect the major agronomic traits (such as plant height, panicle number per plant, grain number per panicle, and seed-setting frequency). Incorporation of the wx mutation into varieties with low initial AAC levels resulted in further reduction in AAC, but without significantly affecting the original, desirable gelatinization traits and amylopectin structure types, suggesting that parents with low initial AAC should be preferred in breeding programs.

Genetic Effects of Indica Lineage Introgression on Amylopectin Chain Length Distribution in Japonica Milled Rice

The fine structure of amylopectin affects rice quality; in particular, the amylopectin chain length distribution (ACLD) in milled rice differs between subspecies of Oryza sativa L. However, the correlation between ACLD and quality trait factors, and the genetic basis of ACLD phenotypic variation, are still unknown. Here, the correlations of ACLD with cooking and eating quality and with the rapid viscosity analysis (RVA) index were studied using chromosome segment substitution lines (CSSLs). Clear variations in ACLD were observed in introgression lines: introgression of indica segments of chromosome 3 and 7 increased the proportion of amylopectin Fa, and another segment of chromosome 3 reduced the proportion of amylopectin Fb2. A segment of chromosome 11 decreased the proportion of amylopectin Fa but increased that of Fb3. Correlation analysis with the RVA index further showed that the breakdown viscosity (BDV) was negatively correlated with the proportion of amylopectin Fb1, Fb2, and Fb3 chains, and positively correlated with Fa. Consistency viscosity (CSV) values were negatively correlated with the proportion of amylopectin Fb1, Fb2, and Fb3 chains. We thus clarified the quality trait factors determined by variation in ACLD, and provide key information for pyramiding inter-subspecific genetic superiority in molecular design breeding for rice quality.

Changes in fine structure of amylopectin and internal structures of starch granules in developing endosperms and culms caused by starch branching enzyme mutations of japonica rice.

BEIIb plays a specific role in determining the structure of amylopectin in rice endosperm, whereas BEIIa plays the similar role in the culm where BEIIb is absent. Cereals have three types of starch branching enzymes (BEs), BEI, BEIIa, and BEIIb. It is widely known that BEIIb is specifically expressed in the endosperm and plays a distinct role in the structure of amylopectin because in its absence the amylopectin type changes to the amylose-extender-type (ae-type) or B-type from the wild-type or A-type and this causes the starch crystalline allomorph to the B-type from the wild-type A-type. This study aimed to clarify the role of BEIIa in the culm where BEIIb is not expressed, by using a be2a mutant in comparison with results with be2b and be1 mutants. The results showed that the amylopectin structure exhibited the B-type in the be2a culm compared with the A-type in the wild-type culm. The starch granules from the be2a culm also showed the B-type like allomorph when examined by X-ray diffraction analysis and optical sum frequency generation spectroscopy. Both amylopectin chain-length profile and starch crystalline properties were found to be the A-type at the very early stage of endosperm development at 4-6days after pollination (DAP) even in the be2b mutant. All these results support a view that in the culm as well as in the endosperm at 4-6 DAP, BEIIa can play the role of BEIIb which has been well documented in maturing endosperm. The possible mechanism as to how BEIIa can play its role is discussed.

The relationship between amylopectin fine structure and the physicochemical properties of starch during potato growth

The aim of this study was to explore the relationship between the structural and functional properties of starch isolated from Atlantic potatoes at different stages of growth without the effect of varieties and growth environment. The molecular size and chain-length distribution of amylopectin significantly varied with growth. The Mw and Mn of amylopectin ranged from 2.976 × 107 to 4.512 × 107 g/mol and 1.275 × 107 to 2.295 × 107 g/mol, respectively, suggested that the polydispersity varied with growth. The average chain length of amylopectin during potato growth showed small but significant changes and ranged from DP 23.59 to 24.73. Overall, Afp chains, Acrystal chains, and B1 chains increased with growth, and B2 and B3 chains decreased with growth. There was wide variation in starch pasting, gelatinization, retrogradation, in vitro starch digestibility, swelling power, solubility, and gel stability properties. Specifically, potato starch harvested at the earliest time had the highest resistant starch content. The variation trend of swelling power and solubility was similar, reached highest value at 42 days, were 20.38 g/g and 8.83%, respectively. Correlation analysis revealed that the physicochemical properties were significantly affected by amylopectin fine structure. The results of this study enhance our understanding of the structure–function relationship of potato starch.

Relationships between supramolecular organization and amylopectin fine structure of quinoa starch

The molecular and granular structure of 9 quinoa starches were investigated and correlated with each other. The molecular structure of quinoa amylopectin and β-limit dextrins studied by high performance anion exchange chromatography suggested that quinoa amylopectin contained a high amount of short chains. The average external chain length (ECL) and average internal chain length (ICL) of the nine quinoa starch studied were 10.7 and 4.90 glucosyl residues, respectively. Small-angle X-ray scattering (SAXS) was applied to study the lamellar structure of quinoa starches. The average thickness of crystalline lamellae (dc) and amorphous lamellae (da) were 6.07 and 2.60 nm, respectively. Correlation analysis suggested that the amounts of short amylopectin chains were positively correlated with dc and they induced heterogeneity in the crystalline lamellae. The amount of long internal chains was positively correlated with da and they could stabilize the double helices in the crystalline lamellae. The correlation analysis based on individual chains suggested that current chain classification might lack a strong structural basis for the supramolecular organization of the granules. The correlations obtained in current research could be explained using the building block backbone model.

Fine structure of amylopectin and relation with physicochemical properties of three coloured potato starches

SummaryIn this study, yellow, red and purple potato starches were selected as the research objects to analyse the fine structure and the relation to the physicochemical properties. Enzymatic hydrolysis and high‐performance anion‐exchange chromatograph were employed to characterise the structure of clusters and φ, β‐limit dextrins. The average degree of polymerisation of clusters from the yellow potato starch was larger (188.57) than in red (91.31) and purple (107.32) potato starch. The molar percentage of fingerprint B chains in yellow, red and purple potato amylopectin were 58.01%, 63.60% and 60.78%, respectively, while major part of short B chains were 15.92%, 17.16% and 16.49%, respectively. The yellow potato amylopectin showed the highest density of branches values indicated that it was more tightly branched. The Pearson correlation coefficients results indicated that the fine structure of amylopectin had significant effects on the physicochemical properties of potato starches, and we can better understanding the differences of the properties among the three potato starches by studying the amylopectin fine structure.

Brassinosteroid receptor mutation influences starch granule size distribution in barley grains

Brassinosteroids (BR) are plant-based steroids which influence several morphogenetic and developmental processes. A barley (Hordeum vulgare L.) genotype Kinai Kyoshinkai-2 (KK-2) carrying the uzu mutation exhibited altered starch granule size distribution. Hybridizing KK-2 with a barley genotype CDC Kendall with bi-modal starch granules produced progeny lines (116, 144 and 168) with almost uni-modal starch granules. Bioassays correlated uzu mutation with defective BR perception. DNA sequence analysis of the BR receptor-1 (BRI-1) gene detected a single-nucleotide A > G substitution at the position 2612 in the kinase domain which resulted in the change of His (CAC) to Arg (CGC) at residue 857 in subdomain IV of the kinase domain of the respective polypeptide. The study focused on the development of barley grain, accumulation of starch and composition influenced by defective BR perception due to the mutation detected in KK-2 and three other barley-breeding lines (116, 144 and 168). Aberrant BRI-1 delayed grain development, amylose synthesis and starch accumulation in the endosperm. The barley breeding lines 116, 144 and 168 carrying the aberrant BRI-1, exhibited altered granule size distribution with significant shift in the diameter maxima, but insignificant differences in amylose concentration. The BRI-1 mutation also altered amylopectin fine structure in both B- and C- type small starch granules, resulting in an increased fraction of short A-type glucan chains (<10 DP) and decreased fraction of B2 chains (25–36 DP) in genotypes carrying the BRI-1 mutation. The results show the influence of BR on barley grain development, starch accumulation, granule size distribution and amylopectin structure.

Effects of amylose and amylopectin fine structure on sugar-snap cookie dough rheology and cookie quality

Starch, the most abundant component of wheat flour, has a significant influence on sugar-snap cookie quality, but the underlying mechanisms are not well understood. In this study, the solvent retention capacity and rheological properties of sugar-snap cookie dough, and cookie physical characteristics and textural properties, were analyzed. Starch molecular structures were measured by size-exclusion chromatography (SEC) and the results parameterized using two biosynthesis-based mathematical models to obtain structure-property correlations. The results show that the viscoelasticity of sugar-snap cookie dough is positively correlated with the length of amylopectin short chains. In addition, the length of amylose short chains is positively correlated with cookie thickness and negatively correlated with its spread ratio. Starch molecular structure shows no influence on cookie hardness and fracturability. Mechanisms are proposed for these results, which could lead to new ways to improve sugar-snap cookie quality by choosing starches with appropriate structural features.

Characterization of eating quality and starch properties of two Wx alleles japonica rice cultivars under different nitrogen treatments

To understand the effect of nitrogen (N) fertilizer on rice (Oryza sativa L.) eating and cooking quality (ECQ). Here, we investigated the ECQ attributes, physicochemical foundation of ECQ, and amylopectin fine structure of two Waxy (Wx) alleles japonica rice cultivars Nanjing 9108 (NJ9108) and Huaidao 5 (HD5) under four N rates (0, 150, 300, and 450 kg ha−1). Sensory and pasting properties of the two cultivars varied depending on N rates. Compared with the control (0 kg ha−1), the overall eating quality and sensory value were significantly decreased under the N rates of 300 and 450 kg ha−1. Further, conventional descriptive analysis showed that the stickiness and retrogradation of cooked rice were significantly decreased. These results indicated that application of N fertilizer seems to affect the texture of cooked rice, causing it to be less sticky, lowering its retrogradation, and consequently reducing its palatability. Results from rapid visco analyzer (RVA) revealed that the peak and breakdown viscosities significantly decreased, while the setback viscosity and peak time increased under the N rates of 300 and 450 kg ha−1. However, no significant difference was observed when the N rate was 150 kg ha−1, indicating that less N fertilization can maintain rice ECQ. As the N rate increasing, protein content increased, whereas apparent amylose content, starch content, and gel consistency almost unchanged. Interestingly, compared with the control, under N treatments, the percentage of short amylopectin branches in NJ9108 was decreased, but increased in HD5, as controlled by amylopectin synthesis-related genes. Notably, SSI and BEIIb were down-regulated in NJ9108, whereas BEIIb was up-regulated in HD5. Thus, the palatability of both rice cultivars was significantly decreased under excessive N fertilization as a consequence of reduced stickiness and retrogradation of the cooked rice, which might have resulted from an elevated protein content and altered amylopectin fine structure. In addition, amylopectin synthesis appeared to be affected by N fertilizer and the genotype of the rice cultivar.

Gelatinization, pasting and retrogradation properties and molecular fine structure of starches from seven cassava cultivars

Genetic diversity in the physicochemical properties and fine structures of seven cassava starches samples was studied. The apparent amylose content ranged from 24.8 to 27.6%. The whole branched starches showed significant differences in average hydrodynamic radius, ranging from 53.35 to 58.45 nm, while debranched starch exhibited differences in degrees of polymerization and height of both amylose and amylopectin peaks. The molecular size of amylose and amylopectin was positively correlated. The amount of short chains fa (6 ≤ X ≤ 12) and fb1 (13 ≤ X ≤ 24) had significant differences among the cultivars. Structure-function relation analysis indicated that the CPV and SB were mainly determined by amylopectin fine structures, BD, PTi and Tp and retrogradation properties were mainly determined by the amylose fine structure, while PTe and To were mainly affected by both amylose and amylopectin fine structures. The current findings will be helpful to improve the understanding cassava starch quality for use in industrial starch applications.

Starch branching enzymes contributing to amylose and amylopectin fine structure in wheat

Starch branching enzymes (SBEs) play a major role in determining starch molecular structure in cereal endosperm. This study investigates how SBEIIs contribute to the chain-length distributions (CLDs) of both amylopectin and amylose, obtained by enzymatic debranching of native starch. Wheat starches with low (37%) to high (93%) amylose content were obtained through altering SBEII in planta. Multiple components were detected in both amylose and amylopectin CLDs. Model fitting of these CLDs reveals a quantitative association between the enzyme activities in amylopectin and amylose. SBEIIa modifies shorter branches (degree of polymerization DP ≲ 12) in amylopectin and longer amylose chains with a CLD peak at ˜3000 DP. SBEIIb acts on longer branches (DP≲ 32) in amylopectin, while its effect on amylose fine structure is not significant. Using both the amylose and amylopectin models to analyze the CLD reveals connections between amylose and amylopectin in wheat starch biosynthesis.

Starch structure-property relations as a function of barley germination times

Two varieties of barley samples were subjected to germination conditions to investigate the underlying mechanisms underpinning changes in molecular structure, chemical compositions and thermal properties of starch during this process. Starch thermal transitions were examined using differential scanning calorimetry, and the molecular fine structure of amylose and amylopectin were determined using size-exclusion chromatography and fluorophore-assisted carbohydrate electrophoresis, respectively. Both amylose and amylopectin chains were hydrolyzed during germination, but a preferential attack of amylopectin chains was observed with concomitant increases of relative amylose content, resulting in increased gelatinization temperatures (onset, peak, conclusion) and reduction in enthalpy change. Amylolytic enzyme activities increased during germination, resulting in decreased starch content. After malting, significant degradation of amylose chains followed by the reduction of gelatinization temperatures was seen. Roasting of pale malts was found to degrade starch and protein whilst completely stopping enzyme activities. The resulting coloured malts had extremely low starch enthalpy change due to the loss of amylopectin crystallinity at high temperature. This study provides insights into starch structural changes of barley throughout malting and roasting, which are determining factors for fermentable sugar production during mashing.

Fine molecular structure and its effects on physicochemical properties of starches in potatoes grown in two locations

Seven potato cultivars were planted at two different locations, E1 (Yiwu) and E2 (Jinhua) to assess the starch molecular structures and physicochemical properties as affected by growing locations. Most of the potato cultivars in E2 presented significantly smaller average whole molecular size (Rh¯) whereas larger average chain length of amylopectin (X¯) along with lower proportion of fa chains (DP 6–12) than those in E1. Significant varietal difference in the physicochemical properties were observed within each location, but only the mean onset temperature in E1 was significantly lower than that in E2. The fine structures (chain-length distribution) of both amylose and amylopectin affected the pasting properties of potato starches based on the starch structure-property correlations. The amylopectin fine structure also determined the thermal and retrogradation properties. Potato starches with more short chains and less long chains of amylopectin displayed lower gelatinization temperature but higher retrogradation.

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.