Isoamylase (Glycogen 6-glucanohydrolase)

Acceptance of new rice genotypes demanded by rice value chain depends on premium value of varieties that match consumer demands of regional preferences. High throughput prediction tools are not available to breeders to classify cooking and eating quality (CEQ) ideotypes and to capture texture of varieties. The pasting properties in combination with starch properties were used to develop two layered models in order to classify the rice varieties into twelve distinct CEQ ideotypes with unique sensory profiles. Classification models developed using random forest method depicted the overall accuracy of 96 %. These CEQ models were found to be robust to predict ideotypes in both Indica and Japonica diversity panels grown under dry and wet seasons and across the years. We conducted random forest modeling using 1.8 million high density SNPs and identified top 1000 SNP features which explained CEQ model classification with the accuracy of 0.81. Furthermore these CEQ models were found to be valuable to predict textural preferences of IRRI breeding lines released during 1960-2013 and mega varieties preferred in South and South East Asia.

Slowly digestible starches have received interest due to their lower increase of postprandial blood glucose and insulin levels and, hence, modification of starches towards slower digestibility has commercial interest. However, chemical characteristics driving enzymatic (digestive) degradation are not fully unraveled. The digestion properties of starches have been linked to their crystalline type, chain length distribution, amylose content or degree of branching, but content and length of relatively long side-chains in amylopectin has not been paid attention to. Therefore, this research focusses on the unique content and length of amylopectin side-chains from conventional and new starch sources (potato, corn, pea, and tulip) correlated to the enzymatic digestion. The rate of hydrolysis was found to be correlated with the crystalline type of starch, as previously suggested, however, the complete hydrolysis of all starches, independent of the crystalline type and source, was shown to be governed by the content of longer amylopectin chains.

This research focused on the structural characteristics of resistant starches (RSs) that were obtained from corn, potato, and sweet potato and esterified by L-malic acid. Further, the unique effect of the degree of substitution (DS) on the crystalline properties was studied. Different starches were allowed to react with 2 M malic acid (pH 1.5) for 12 h at 130°C. The shapes of the granules and the Maltese-cross shapes of samples were maintained and visible under an optical microscope. The FT-IR spectrum displayed evident carbonyl peaks at 1740 cm⁻¹, and the onset temperature (T_o) and gelatinization enthalpy (∆H) gradually decreased as DS increased. The malic acid-treated starches exhibited an increased RS content compared to those of the control. The RS contents of potato, sweet potato, and corn, which were 65.5%, 70.0%, and 89.8% in the uncooked MT-samples, decreased to 57.3%, 63.8%, and 86.7% in the cooked MT-samples, respectively, and exhibited high heat stability; corn starch yielded the highest RS among them. The thermal and malic acid treatments resulted in the partial hydrolysis and rearrangement of the helix structure of crystalline area, which was affected by esterification. The result revealed that the RS content increased as that of DS escalated.

Rice milling is a very common process to improve rice edible quality. In this study, the effects of the degree of milling (DOM) on starch leaching characteristics and its relation to rice stickiness were investigated. As DOM increased, the whiteness of rice grains and rice stickiness increase, while the protein and lipid content of rice grains decreases. By examining the starch leaching characteristics during rice cooking, it shows that i) increasing DOM significantly increases the total solids and leached starch content while decreases the leached protein content; ii) the molecular size and chain-length distribution (CLD) of leached starch are not significantly varied between rice samples with different DOM, but are significantly different from that of native starch; iii) a significant correlation between rice stickiness and leached amylopectin amount is established; iv) the molecular mechanism for the increased stickiness as affected by DOM is also proposed. The improved understanding of rice processing obtained in this study may allow a better control of rice eating quality.

This study investigated effects of longan seed polyphenols (LSPs) on the structure and digestion properties of starch, and discussed the interaction mechanism between starch and LSPs. The results showed cooking with 20 % LSPs did not change amylopectin chain length distribution of normal maize starch, however, the amylose content was reduced from 21.60 to 14.03 %. This suggests LSPs may interact with starch via non-covalent bond. Isothermal titration microcalorimetry and XRD results confirmed the existence of non-covalent interaction, and indicated that LSPs may enter the hydrophobic cavity of amylose, forming V-type inclusion complex. LSPs did not affect gelatinization temperatures of maize starch, whereas 20 % LSPs decreased the enthalpy change by about 26 %. The digestion results indicate significant inhibition effect of LSPs on the digestion of cooked starch, attributing to the interaction of LSPs with starch. These suggest potential applications of LSPs as functional ingredients in modulating postprandial glycemic response of starchy food.

Native rice starch is a source of slowly digestible starch in e.g. low-moisture baked products, while the molecular and lamellar/crystalline structure giving rise to this low-digestibility property is still largely unknown. In this study, the in vitro digestion kinetics of 11 rice starches with a wide range of amylose content were investigated. Applying the logarithm of slope (LOS) plot to the starch digestograms suggested that only a single first-order kinetics phase existed. More importantly, results for the first time showed that rice starches with shorter amylopectin short chains (DP 10-26) had more perfectly aligned crystalline lamellae and much slower digestion rates than the other starches. Interestingly, no correlations were found between the starch lamellar thicknesses with its digestion rate. It suggests that lamellar perfection plays a dominant role in the determination of native starch digestibility. Furthermore, starches with relatively shorter amylose short and medium chains showed a significantly higher amount of V-type amylose-lipid complex, and smaller maximum digestion extent. These results could help the rice industry develop a new generation of rice products with slower starch digestion rate and more desirable nutritional values.

In this study, three rice starches with different amylose content were isolated from three rice varieties, followed by chain elongation using amylosucrase from Neisseria polysaccharea (NpAS). Herein, the chain elongation could induce the starch precipitation during the enzymatic reaction. With a higher content of amylose, an earlier precipitation occurred, leading to the decrease of transglycosylation efficiency of the enzyme. Results from X-ray diffraction and thermal property analysis indicated that the crystalline structure of the NpAS-modified starches largely formed at the precipitation point. The elongation at the surface of the starch precipitates, however, would not enable the formation of crystallites. In addition, the amylose might participate in the recrystallization of starch molecules. Digestion kinetics revealed that all of the NpAS-modified starches contained rapidly and slowly digested fractions, the latter of which had a digestion rate being dependent on the structural stability of crystallites (length of double helices). These findings may provide an efficient way to produce chain-elongated starches with desirable functionality and digestibility.

Starch from bananas/plantains, belonging to the genus Musa spp, is gaining prominence given its great potential as a healthy food ingredient made from an inexpensive raw material. Recent works highlight the outstanding potential of Musa starch to develop enzyme-resistant structures upon retrogradation. However, despite the wide variety of Musa cultivars (due to both natural mutation and breeding selection), there is no comparative investigation of the starch molecular structure from the most commonly cultivated Musa genotypes. In this work, the starch from six Musa cultivars harvested during the same growing season from the same parcel, was purified and analyzed for amylose ratio, amylose chain length distribution, and amylopectin unit and internal chain length distribution. Results showed significant differences between the fine structure of all Musa amylopectin molecules, which were structurally categorized as type 4 (consisting of a high number of B3-chains, few BS- and B_fp-chains, and low S:L and BS:BL ratios). Moreover, the different Musa starches exhibited dramatic differences in amylose ratio (17.7-27.6%), amylose branching degree (as evidenced by differences in the population of short chains of approximately 260 glucose units, GU) and a shorter average length (approximately 1000 GU) of the population of long amylose unit chains. Remarkably, these differences in amylose structure resulted in the cultivar Manzano (Musa AAB, silk subgroup) to possess a dramatically lower extension of in vitro starch digestion (C₉₀ = 4.70%) than the rest of the cultivars (C₉₀ = 17-18%) after full gelatinization and retrogradation for 7 days.

Waxy maize starch was debranched using isoamylase and simultaneously crystallized in the same batch at 50°C for 7 days. The crystallized starch dextrins were isolated as precipitates and characterized for chain length distribution, crystallinity, thermal transition property, morphology, and in vitro digestibility. The long B chains released from the amylopectin crystallized more readily than the short A chains. The dextrins exhibited a typical A-type crystalline arrangement with high thermal stability. The recovery yield from the waxy maize starch was 70.4% after 7 days. The amount of resistant starch (RS) in the crystallized dextrin treated for 7 days was 83% (uncooked) or 46% (cooked). The simultaneous debranching and crystallization process was effective for the formation of dextrin crystals from waxy maize starch with thermal stability and good resistance to digestive enzymes.

We describe a method for permitting efficient modification by transglucosidase (TGA), from glycoside hydrolase family 31 (GH31), sequentially after the pre-treatment by maltogenic α-amylases (MA) from GH13. TGA treatment without MA pre-treatment had negligible effects on native starch, while TGA treatment with MA pre-treatment resulted in porous granules and increased permeability to enzymes. MA→TGA treatments lead to decreased molecular size of amylopectin molecules, increased α-1,6 branching, and increased amounts of amylopectin chains with the degree of polymerization (DP)<10 and decreased amounts of DP 10-28 after debranching. Wide-angle X-ray scattering (WAXS) data showed a general decrease in crystallinity except for a long term (20 h) TGA post-treatment which increased the relative crystallinity back to normal. MA→TGA treatment significantly lowered the starch retrogradation of starch and retarded the increase of storage- and loss moduli during storage. This work demonstrates the potential of sequential addition of starch active enzymes to obtain granular starch with improved functionality.

A neutral polysaccharide (NPP) from peanut sediment of aqueous extraction process was purified via anion-exchange and gel-filtration chromatography. The weight-average molecular weight and polydispersity index were 3.36 × 10⁴ Da and 1.06. Composition of glucose (82.66 %, molar percentage) and arabinose (17.34 %) suggested an arabinoglucan structure. Multiple medium-length chains consisting of many 1,4-linked α-Glcp and a few 1,5-linked α-Araf maintained the main chain structure. The backbone was substituted at O-6 and O-3 positions, attached by side chains consisting of two to six α-Glcp and terminated with Araf and Glcp. Degree of branching was 42.50 %. Aggregates formed in NPP aqueous solution. They were eliminated by DMSO combining with sonication. Consequently, the average radius of gyration (R_g), hydrodynamic radius (R_h), and R_g/R_h ratio were 17.0 nm, 5.8 nm and 2.93, respectively, indicating extended rigid chain conformation. The backbone substituted at O-3 and short branching chains probably together induced this conformation.

This study investigated the effects of acid degradation of amylopectin on the structure, pasting, and rheological properties of waxy maize starch. It is found that: 1) the amount of amylopectin short-chains with degree of polymerization (DP) ~15-50 increased while that of amylopectin long-chains with DP ~50-200 decreased by acid hydrolysis; 2) acid hydrolysis produced smaller amylopectin molecules with a narrower size distribution; 3) acid hydrolysis had a minor effect on the crystalline and granular structures of native starch; 4) the pasting viscosity of acid hydrolyzed starch during heating and the consistency coefficient, K, of starch gels increased, whereas the flow behavior index, n, decreased. Correlation analysis was used to clarify the molecular causes for the variations of pasting and rheological properties of acid hydrolyzed starch.

This study uses sunflower hulls, a by-product from the sunflower snack industry, to recover both, valuable phenolic compounds and cellulose fibers, for the production of antioxidant reinforced starch films as potential food packaging material. The phenolic extract provided antioxidant properties to the films with EC₅₀ values of 89 mg film/mg DPPH. The cellulose fibers reinforced the starch films with a threefold increase in Young´s modulus. Furthermore, citric acid was added to induce cross-linking of the starch polymers and improve film integrity. The addition of citric acid induced both, starch polymer hydrolysis and cross-linking, seen in a shift in chain-length distribution after debranching with iso-amylase. This is the first study that focuses on a three-principle approach to improve edible starch films, and follows UN goals on sustainability to reduce waste and increase value in by-products as a step forward to functionalize packaging material.

Peach palm fruit mesocarp (Bactris gasipaes var. gasipaes) is already consumed in the Northern region of Brazil, after its cooking and is known as a source of starch and carotenoids and like all fruits it has low storage stability. This work characterized the starch extracted from the mesocarp of peach palm fruit using with water in terms of its physical and chemical properties. The SEM micrographs show that starch presented bimodal distribution (size 3.9-10.4 µm), while the smaller granules had a smooth surface and an oval or conical shape, the larger granules were spherical with holes and cracks on the surface. The starch presented low amylose content (<20%) and amylopectin branch chain length distribution with the absence of a shoulder, which is suggestive of perfect crystalline structure, and a higher proportion of medium chains (DP 13-24), despite the large number of short chains (DP 6-12), and on average DP 21. X-ray diffraction showed a mixture of polymorphs A and B, which can be considered C-type crystalline pattern, which is indicative of being a slow digestible starch. Through paste viscosity results, by RVA, we can observe low values for thermal and pasting properties, suggesting greater homogeneity of crystals. Also, due to interaction with lipids originally present (2.69%), the starch showed lower retrogradation rate (22.64%), which resulted in a weak gel after 24 h of storage. As a product with greater storage stability, peach palm fruit starch, extracted for the purpose of promoting its regional use, has shown that it can be used in products where slow and smooth retrogradation is desired, such as in breads, soups, chowder and porridges, without the use of emulsifiers or fat.

We performed a dual-modification of starch via debranching and graft copolymerization to improve its adhesion to fibers. We synthesized the partly debranched starch-g-poly(2-acryloyloxyethyl trimethyl ammonium chloride) (PDS-g-PATAC) using horseradish peroxidase in the presence of hydrogen peroxide and acetylacetone. PDSs of different molecular structures were prepared by debranching waxy cornstarch for different periods of time. With increasing debranching time, the degree of hydrolysis of PDS increased from 0.85 % (10 min) to 1.13 % (30 min), while the degree of branching decreased from 8.37 % to 7.99 %. Fourier transform infrared analysis confirmed that ATAC units had been successfully grafted onto the starch (debranched or not debranched). The degree of substitution (DS) and grafting ratio (GR) of the PDS-g-PATACs were characterized by ¹H nuclear magnetic resonance. The DS and GR of grafted starches positively related with debranching time. Thermogravimetry-differential thermogravimetry analysis showed that grafted starch had lower thermal stability than ungrafted starch. The adhesion of PDS-g-PATAC to polyester/cotton yarns was evaluated. The PATAC chains grafted onto the starch enhanced the adhesion of starch to polyester/cotton yarn. The grafted starch produced from the copolymerization of PDS (after debranching for 10 min), exhibited the strongest adhesion to polyester/cotton yarn with a resulting tensile strength of 98.20 N.

The objective of this study was to investigate the structural and physicochemical properties of starch from seven sweet potato cultivars (Shinyulmi, Sinjami, Hogammi, Jeonmi, Jinyulmi, Juhwangmi, and Pungwonmi). Jeonmi and Jinyulmi had amylose contents of 40.04% and 37.39%, respectively, whereas Juhwangmi and Pungwonmihad amylose contents of 30.95% and 32.37%, respectively. As a result of amylopectin polymerization, the seven cultivars were found to have high (>48%) contents of the degree of polymerization (DP) 13~24 fraction, whereas the DP≥37 fraction content was <3.45%. The level of resistant starch was highest in Jeonmi (>30%) and lowest in Pungwonmi (<5%). The in vitro digestibility of Pungwonmi was greater than that of the other cultivars. Starch X-ray patterns did not differ among the cultivars. The results of this study provide useful information for the food industry regarding the application of sweet potato starches.

Starch granule-associated proteins (SGAPs) include granule-surface proteins (SGSPs) and granule-channel proteins (SGCPs). To investigate impacts of SGAPs on amyloglucosidase (AMG) hydrolysis, waxy and non-waxy rice starches had their SGCPs or SGAPs removed. Removal of SGAPs or SGCPs did not affect morphology and amylopectin chain distribution but decreased relative crystallinity. Removal of SGAPs increased the digestion rate, AMG binding ability and pore diameter of hydrolyzed granules, and accelerated changes in relative crystallinity and destruction of crystalline region on hydrolysis. However, after removing SGCPs, AMG only bound to surface and attacked of the fingerprint of protein bodies on granules, with decreased hydrolysis rate. The degree of change in hydrolysis rate was not determined by SGCPs content of rice starch. These results implied that SGCPs had a more dominant role in AMG hydrolysis of rice starch than did SGSPs. This study provides novel information about the role of SGAPs in AMG hydrolysis mechanisms.

A structure-digestion model is proposed to explain the formation of α-amylase-slowly digestible structures during amylopectin retrogradation. Maize and potato (normal and waxy) and banana starch (normal and purified amylopectin through alcohol precipitation), were analyzed for amylose ratio and size (HPSEC) and amylopectin unit- and internal-chain length distribution (HPAEC). Banana amylopectin (BA), like waxy potato (WP), exhibited a larger number of B3-chains, fewer BS- and B_fp-chains and lower S:L and BS:BL ratios than maize, categorizing BA structurally as type-4. WP exhibited a significantly greater tendency to form double helices (DSC and ¹³C-NMR) than BA, which was attributed to its higher internal chain length (ICL) and fewer DP6−12-chains. However, retrograded BA was remarkably more resistant to digestion than WP. Lower number of phosphorylated B-chains, more S- and B_fp-chains and shorter ICL, were suggested to result in α-amylase-slowly digestible structures through further lateral packing of double helices (suggested by thermo-rheology) in type-4 amylopectins.

The physicochemical properties and starch fine structure change during paddy storage, which may be beneficial for the qualities of rice-based products such as fresh rice noodles (FRN). In this study, the textural properties of FRN prepared from polished rice of six indica paddy cultivars after different storage period at 36 ± 2°C and 85% ± 5% relative humidity were investigated. The pasting properties, solubility (S), swelling power (SP) of rice flour and morphology of the starch granules, chain length distribution of amylopectin were analyzed to determine factors that impact the texture of FRN. The chewiness and springiness of FRN increased, and the adhesiveness decreased with increase in storage duration. The setback (SB) and pasting temperature (PT) increased while the S and SP decreased with extension of storage duration (0-90 days). Chain length distributions of amylopectin also changed. Pearson correlation analysis revealed that cool paste viscosity (CPV) of indica rice flour, short chain (DP 6-12) and long chain (DP 37-60) of amylopectin were the most correlated parameters with adhesiveness of FRN. Our results may provide insights for selecting raw materials for production of high-quality FRN.

To develop a 1,4-α-glucan branching enzyme (BE) without homology to known allergens, the glgB gene from Bifidobacterium longum was overexpressed under the control of BLMA promoter in Escherichia coli. B. longum BE (BlBE) had a molecular weight of 86.1 kDa and a specific activity of more than 18.5 U/mg protein at 25-35°C and pH 5.5-7.0, and exhibited 30% of the maximum activity at 10°C. The cold-active BlBE preferred to transfer maltohexaose and introduced DP 4-36 branches into amylose. BlBE also increased the proportion of DP 2-10 branches in amylopectin and decreased its Mw from 1.39 × 10⁶ to 1.16 × 10⁵ g/mol. As the BlBE concentration increased from 0.0 to 0.5 U/mg substrate, the retrogradation enthalpy of BlBE-modified wheat starch decreased from 4.50 to 1.83 J/g (p < 0.05) at day 14 and the slowly digestible starch content increased from 2.10% to 17.39% (p < 0.05).