Isoamylase (Glycogen 6-glucanohydrolase)

The effect of heat-moisture treatment on structural, physicochemical, and rheological characteristics of arrowroot starch was investigated. Heat-moisture treatment was performed with starch samples conditioned to 28% moisture at 10°C for 2, 4, 8, and 16 h. Structural and physicochemical characterization of native and modified starches, as well as rheological assays with gels of native and 4 h modified starches subjected to acid and sterilization stresses were performed. Arrowroot starch had 23.1% of amylose and a C_A-type crystalline pattern that changed over the treatment time to A-type. Modified starches had higher pasting temperature and lower peak viscosity while breakdown viscosity practically disappeared, independently of the treatment time. Gelatinization temperature and crystallinity increased, while enthalpy, swelling power, and solubility decreased with the treatment. Gels from modified starches, independently of the stress conditions, were found to have more stable apparent viscosities and higher G′ and G″ than gels from native starch. Heat-moisture treatment caused a reorganization of starch chains that increased molecular interactions. This increase resulted in higher paste stability and strengthened gels that showed higher resistance to shearing and heat, even after acid or sterilization conditions. A treatment time of 4 h was enough to deeply changing the physicochemical properties of starch.

Starch is the principal component of rice that affects its cooking and nutritional quality. This study investigated molecular and thermal properties of starches isolated from seven Africa rice accessions (ARAs) in comparison with two commonly produced Asian rice varieties (ARVs) and a developed cross (sativa × glaberrima) variety (NERICA 4). All starch granules were polyhedral and tightly packed with size distribution ranging from 2-22 µm and displayed type-A X-ray diffraction pattern. ARAs starch granules had higher ratio of absorbance to scattering when exposed to iodine vapor exhibiting greater flexibility and availability of glucan chains to form complexes with iodine as compared to ARVs. The enthalpies of starch gelatinization (15.1-15.8 J/g) and retrograded gel melting (9.2-10.8 J/g) were higher in ARAs than in NERICA 4 (14.5 and 9.2 J/g, respectively) and ARVs, (13.3-14.3 and 6.4-7.3 J/g, respectively) possibly due to their higher amylose content and longer chains. Significant (p < 0.05) differences in peak, trough, final, breakdown, and setback viscosities were also observed among the starches with Koshihikari Asian rice having the highest peak viscosity (310 RVU). These differences in molecular structure and thermal properties between the ARAs and ARVs are likely to influence the cooking and eating quality of the ARAs.

Pinhão seed is an unconventional source of starch and the pines grow up in native forests of southern Latin America. In this study, pinhão starch was adjusted at 15, 20 and 25% moisture content and heated to 100, 110 and 120°C for 1 h. A decrease in λ max (starch/iodine complex) was observed as a result of increase in temperature and moisture content of HMT. The ratio of crystalline to amorphous phase in pinhão starch was determined via Fourier transform infra red by taking 1045/1022 band ratio. A decrease in crystallinity occurred as a result of HMT. Polarised light microscopy indicated a loss of birefringence of starch granules under 120°C at 25% moisture content. Granule size distribution was further confirmed via scanning electron microscopy which showed the HMT effects. These results increased the understanding on molecular and structural properties of HMT pinhão starch and broadened its food and nonfood industrial applications.

A cold-active α-amylase, Amy_13C6, identified by a functional metagenomics approach was expressed in Escherichia coli and purified to homogeneity. Sequence analysis showed that the Amy_13C6 amylase was similar to α-amylases from the class Clostridia and revealed classical characteristics of cold-adapted enzymes, as did comparison of the kinetic parameters K _m and K _cat to a mesophilic α-amylase. Amy_13C6 was shown to be heat-labile. Temperature optimum was at 10–15°C, and more than 70 % of the relative activity was retained at 1°C. The pH optimum of Amy_13C6 was at pH 8–9, and the enzyme displayed activity in two commercial detergents tested, suggesting that the Amy_13C6 α-amylase may be useful as a detergent enzyme in environmentally friendly, low-temperature laundry processes.

The effects of chain distribution, concentration, temperature and hydrothermal treatments on the recrystallization behavior and formation of resistant starch (RS) were investigated. Waxy and normal rice starches were debranched at 10 and 21% w/w solid concentrations, incubated at 25 or 50°C, and further subjected to annealing or heat moisture treatment (HMT) to enhance RS formation. The crystallization at 25°C favored the formation of the B-type structure, whereas crystallization at 50°C led to the A-type structure with a higher melting temperature (100-120°C) and a higher RS content (52%). All incubated samples showed an increase in RS content after subsequent hydrothermal treatments. The sample incubated at a high temperature contained the highest RS content (74.5%) after HMT with larger/perfect crystallites. These results suggested that the RS formation could be manipulated by crystallization conditions and improved by hydrothermal treatments which are dependent on the initial crystalline perfection.

The impact of starch granule structure on amylose (AM) and amylopectin (AP) chain reactivities was investigated over a 24 h period in a model reaction system utilizing a fluorescent probe [DTAF, 5-(4,6-dichlorotriazinyl)aminofluorescein] as a reagent. For various reaction time intervals (0.5, 4, 12, or 24 h), molecular reactivities of debranched starch derivatives were assessed via size-exclusion chromatography equipped with refractive index and fluorescence detection. For all starch chain fractions, the initial rate of derivatization (0-0.5 h) was rapid, but decreased thereafter. Starch chain reactivities followed the general order: AP long chains≥ AM, AP medium chains > AP short chains, though both AM and AP long chain reactivities were somewhat impacted by the high relative reactivity of starch chains eluting in the AM/AP long chain boundary region. Varied starch chain reactivities were attributed to their relative locale within the granule, corroborating the impact of granule architecture on molecular-level reaction patterns of starch chains.

Granular normal maize starch was reacted with sodium trimetaphosphate in deionized water (_H2O), aqueous sodium sulfate solution (MSNa2SO4), aqueous ethanol (MS_ethanol) or aqueous acetone (MS_acetone ) under otherwise identical reaction conditions. Analysis of the resultant starches by Rapid Visco Analysis (RVA) showed that the starch was cross-linked to a higher degree in aqueous ethanol or aqueous acetone than in water or sodium sulfate solution, and with minimal starch leaching. While MS_acetone and MS_ethanol had incorporated similar levels of phosphorous, RVA analysis and microscopic analysis showed that MS_acetone granules were more effectively stabilized by cross-linking than MS_ethanol granules. Cross-linking in aqueous acetone is believed to either contain the greater numbers of distarch monophosphate (versus monostarch monophosphate), or occur more intensively at the granule outer layers than that in aqueous ethanol and, at the same time, to account for the greater granular strength of MS_ethanol than that of MS_acetone .

The objective of this study was to investigate the structural characteristics of slowly digestible starch (SDS) and resistant starch (RS) fractions isolated from heat–moisture treated waxy potato starch. The waxy potato starch with 25.7% moisture content was heated at 120°C for 5.3 h. Scanning electron micrographs of the cross sections of RS and SDS + RS fractions revealed a growth ring structure. The branch chain-length distribution of debranched amylopectin from the RS fraction had a higher proportion of long chains (DP ≥ 37) than the SDS + RS fraction. The X-ray diffraction intensities of RS and SDS + RS fractions were increased compared to the control. The SDS + RS fraction showed a lower gelatinization enthalpy than the control while the RS fraction had a higher value than the SDS + RS fraction. In this study we showed the RS fraction is composed mainly of crystalline structure and the SDS fraction consists of weak crystallites and amorphous regions.

Starch provides our main dietary caloric intake and over-consumption of starch-containing foods results in escalating life-style disease including diabetes. By increasing the content of α-1,6 branch points in starch, digestibility by human amylolytic enzymes is expected to be retarded. Aiming at generating a soluble and slowly digestible starch by increasing the content and changing the relative positioning of the branch points in the starch molecules, we treated cassava starch with amylomaltase (AM) and branching enzyme (BE). We performed a detailed molecular analysis of the products including amylopectin chain length distribution, content of α-1,6 glucosidic linkages, absolute molecular weight distribution and digestibility. Step-by-step enzyme catalysis was the most efficient treatment, and it generated branch structures even more extreme than those of glycogen. All AM- and BE-treated samples showed increased resistance to degradation by porcine pancreatic α-amylase and glucoamylase as compared to cassava starch. The amylolytic products showed chain lengths and branching patterns similar to the products obtained from glycogen. Our data demonstrate that combinatorial enzyme catalysis provides a strategy to generate potential novel soluble α-glucan ingredients with low dietary digestibility assets.

We determined the relationships among the structural properties, in vitro digestibility, and genetic factors in starches of 14 rice cultivars. Weight-based chain-length distributions in amylopectin ranged from 18.07% to 24.71% (fa, DP 6-12), 45.01% to 55.67% (fb₁, DP 13-24), 12.72% to 14.05% (fb₂, DP 25-36), and 10.80 to 20.72% (fb₃, DP > 36), respectively. The contents of rapidly digestible starch (RDS), slowly digestible starch (SDS), and resistant starch (RS) ranged from 78.5% to 87.5%, 1.2% to 6.0%, and 10.1% to 18.0%, respectively. AAC was negatively correlated with RDS content but positively correlated with RS content in rice starch. The proportion of short chains in amylopectin, i.e. the amount of fraction IIa (FrIIa) fractionated by gel permeation chromatography (GPC), was positively correlated with RDS. Starch synthase IIa (SSIIa) gene controlled the degree of crystallinity, the amount of fa chains of amylopectin. SSIIIa gene controlled the amount of fb1 chains. Wx gene controlled the FrI, FrIIa, RDS, and RS. Starch debranching enzyme isoamylase II (ISA2) gene also controlled the RDS, which may suggest that RDS was also affected by amylopectin structure, although no correlation between them was found. This study indicated that genetics (i.e., starch biosynthesis related genes) controlled the structural properties of starch, and both amylose content and amylopectin fine structure determined functional properties of rice starch (i.e., the digestion), each in a different way. Understanding the “genetics-structure-function” relationships in rice starches will assist plant breeders and food processors in developing new rice varieties and functional foods.

Soluble starch polymers are shown to enhance the lubrication of ionic liquid–water solvent mixtures in low-pressure tribological contacts between hydrophobic substrates. A fraction of starch polymers become highly soluble in 1-ethyl-3-methylimidazolium acetate (EMIMAc)-water solvents with ionic liquid fraction ≥60 wt%. In 65 wt% EMIMAc, a small amount of soluble starch (0.33 wt%) reduces the boundary friction coefficient by up to a third in comparison to that of the solvent. This low-friction is associated with a nanometre thick film (ca. 2 nm) formed from the amylose fraction of the starch. In addition, under conditions where there is a mixture of insoluble starch particles and solubilised starch polymers, it is found that the presence of dissolved amylose enhances the lubrication of starch suspensions between roughened substrates. These findings open up the possibility of utilising starch biopolymers, as well as other hydrocolloids, for enhancing the performance of ionic liquid lubricants.

High salivary amylase activity is associated with improved glycemic homeostasis in humans. Therefore, high salivary amylase activity is associated with greater digestion of starch. However, it is unclear if the structures of the hydrolysates from different individuals with different salivary amylase activity are the same. To test this, cooked starch (1:2 starch/water ratio) was treated with saliva from six participants at equal activity and conditions mimicking oral digestion. Salivary amylase activities ranged from 470 × 10³ to 118 × 10³ U/mL among the participants. The composition of the hydrolysates was characterised by gel-permeation chromatography. All samples gave rise to different and complex mixtures of hydrolysates with different breakdown structures. While saliva from participant 2 (high amylase activity) greatly reduced the high MW fraction, the saliva from participant 6 (low amylase activity) more extensively hydrolysed the starch to small MW fractions of oligosaccharides. These results show that different starch hydrolysates are produced during oral digestion by saliva from different individuals. Further research is therefore needed to understand if hydrolysate structure, rather than level of amylase activity, impacts glucose homeostasis.

Rice plants are known to accumulate starch in leaf sheaths and culms, and in some cultivars significant amounts of starch are present at the mature stage. This can be considered as potential feedstock for the recovery of fermentable sugars. We isolated starches from the culms of cultivars Yumeaoba, Koshihikari, and Leafstar to investigate their structural and physical features. Yumeaoba culm starch contained 20.2% amylose, whereas Koshihikari and Leafstar contained 25.8% and 25.2%. Yumeaoba culm starch was found by chain-length distribution analysis to contain higher amounts of short chains, resulting in lower gelatinization temperature by 7°C, as compared to Koshihikari and Leafstar. Consequently, the rate of enzymatic hydrolysis of Yumeaoba culm starches reached maximum at a lower temperature than Leafstar. Rice culm starch, with a lower gelatinization temperature, can provide an advantageous material for feedstock for bioethanol production in terms of energy conservation.

Inhibition of plant growth by Trinexapac-ethyl, TE, a gibberellin-biosynthesis inhibitor, can produce a shorter stemmed plant, requiring less nutrients and water to grow, while maintaining grain yield. Although TE and other plant growth regulators are commonly used in grain crops, their effects on starch biosynthesis in the grains have not been systematically examined. The changes in the structural and functional properties of starch in grains harvested from TE-treated sorghum (Sorghum bicolor (L.) Moench) were examined, and the results compared with those from the untreated controls. TE treatment had little or no effects on the molecular structures of starch, starch granule morphology, and starch and amylose contents, but increased the protein content of the grains significantly. Consistent with the lack of change in the molecular structure, there were no significant effects on the thermal properties of the starch. The pasting properties of TE-treated sorghum flours, however, showed lower peak viscosity, trough, and final viscosity, which were attributed to their higher protein contents. The TE treatment thus does not have an appreciable effect on the biosynthesis of starch during grain development in sorghum.

Background: Two distinct starch branching enzyme (SBE) isoforms predate the divergence of monocots and dicots and have been conserved in plants since then. This strongly suggests that both SBEI and SBEII provide unique selective advantages to plants. However, no phenotype for the SBEI mutation, sbe1a, had been previously observed. To explore this incongruity the objective of the present work was to characterize functional and molecular phenotypes of both sbe1a and wild-type (Wt) in the W64A maize inbred line. Results: Endosperm starch granules from the sbe1a mutant were more resistant to digestion by pancreatic α-amylase, and the sbe1a mutant starch had an altered branching pattern for amylopectin and amylose. When kernels were germinated, the sbe1a mutant was associated with shorter coleoptile length and higher residual starch content, suggesting that less efficient starch utilization may have impaired growth during germination. Conclusions: The present report documents for the first time a molecular phenotype due to the absence of SBEI, and suggests strongly that it is associated with altered physiological function of the starch in vivo. We believe that these results provide a plausible rationale for the conservation of SBEI in plants in both monocots and dicots, as greater seedling vigor would provide an important survival advantage when resources are limited.

Starches were isolated and characterised from 10 potato cultivars grown under the same conditions (with a commercial starch for reference). The chemical composition revealed some differences amongst the starches with protein ranging from 0.30% to 0.34%, amylose 25.2% to 29.1% and phosphorus 52.6–66.2 mg 100 g^-1. High performance size-exclusion chromatography (HPSEC) fractionation of isoamylase debranched amylopectin showed that the amylopectin molecules were less branched and consisted of more B1, but less A-chains, than cereal starches. Gelatinisation onset (T>_o), peak (T>_p) and conclusion (T>_c) temperatures of the native potato starches ranged from 58.7 to 62.5°C, 62.5 to 66.1°C and 68.7 to 72.3°C, respectively, whilst the gelatinisation enthalpies ranged from 15.1 to 18.4 J g^-1. The gelatinisation temperatures of the starches increased in common with the amounts of short and intermediate sized amylopectin chains. The ¹³ C magic angle spinning nuclear magnetic resonance (¹³C CP-MAS NMR) and wide angle X-ray diffraction (XRD) data (30.6% ± 0.22% crystallinity on average) showed little variance amongst the samples. Particle sizing results, however, revealed more variance (20.6–30.9 µm mean diameter). Overall, these data reveal the subtleties of cultivar specific variation against a background of constant environmental conditions.

Polydextrose (Litesse^®) provides physiological effects consistent with dietary fiber. However, AOAC methods for measuring total dietary fiber (TDF) in foods include an ethanol precipitation step in which polydextrose and similar carbohydrates are discarded and therefore not quantitated. This study describes a method developed to quantitate polydextrose in foods. The new method includes water extraction, centrifugal ultrafiltration, multienzyme hydrolysis, and anion exchange chromatography with electrochemical detection. Six foods were prepared with 4 levels of polydextrose to test the ruggedness of the method. Internal validation demonstrated the ruggedness of the method with recoveries ranging from 83 to 104% with an average of 95% (n = 24) and relative standard deviation of recoveries ranging from 0.7 to 13% with an average of 3.3% (n = 24). The value is added to that obtained for dietary fiber content of foods using the AOAC methods, to determine the TDF content of the food.

Scanning electron microscopy (SEM) and X-ray diffraction (XRD) were employed to investigate degradation patterns of native starch granules from wheat (Triticum aestivum L.) by different starch-degrading enzymes. The starches examined were from a waxy wheat and four varieties with slightly elevated amylose content, but with different functional properties. Differences in the digestion patterns after partial α-amylolysis of starch granules were noted between the starches. The waxy starch seemed to be degraded by endocorrosion, whereas the amylose-rich starches followed a slower mode of hydrolysis starting from the granular surface. X-ray diffractograms of the amylose-rich starches were not significantly altered by 2 h of α-amylolysis, whereas partial hydrolysis of the waxy starch decreased scattering intensity at higher 2θ°angles, consistent with a different mode of attack by α-amylase in the initial digestion stages of granules of waxy and amylose-rich starches. We propose these differences are due to the combined effects of the change in packing density and partial preference for hydrolysis of amorphous material. The native starch granules were also attacked by beta-amylase, isoamylase and amyloglucosidase, which indicates that α-amylase is not the only starch-degrading enzyme that is able to initiate starch hydrolysis of native granules.

Four corn types with different doses of mutant amylose-extender (ae) and floury-1 (fl1) alleles, in the endosperm, including no. 1, aeaeae; no. 2, fl1fl1fl1; no. 3, aeaefl1; and no. 4, fl1fl1ae, were developed for use in making Hispanic food products with high resistant starch (RS) content. The RS percentages in the native starch (NS) of 1−4 were 55.2, 1.1, 5.7, and 1.1%, respectively. All NS were evaluated for pasting properties with a rapid viscoanalyzer (RVA) and for thermal properties with a differential scanning calorimeter (DSC). NS 1 had a low peak viscosity (PV) caused by incomplete gelatinization, whereas NS 3 had the greatest PV and breakdown of all four starch types. On the DSC, NS 2 had the lowest onset temperature and greatest enthalpy. NS 1 and 3 had similar onset and peak temperatures, both higher than those of NS 2 and 4. The gel strength of NS heated with a RVA was evaluated by using a texture analyzer immediately after RVA heating (fresh, RVA-F) and after the gel had been stored at 4°C for 10 days (retrograded, RVA-R). NS 1 gel was watery and had the lowest strength (30 g) among starch gel types. NS 3 gel, although exhibiting syneresis, had greater gel strength than NS 2 and 4. The structures of the NS, the RS isolated from the NS (RS-NS), the RS isolated from RVA-F (RS-RVA-F), and the RS isolated from RVA-R (RS-RVA-R) were evaluated by using size exclusion chromatography. NS 1 had a greater percentage of amylose (AM) (58.3%) than the other NS (20.4−26.8%). The RS from all NS types (RS-NS) had a lower percentage of amylopectin (AP) and a greater percentage of low molecular weight (MW) AM than was present in the original NS materials. The RS-RVA-R from all starches had no AP or high MW AM. The percentages of longer chain lengths (DP 35−60) of NS were greater in 1 and 3 than in 2 and 4, and the percentages of smaller chain lengths (DP 10−20) were greater in 2 and 4 than in 1 and 3. In general, NS 3 seemed to have inherited some pasting, thermal, and structural characteristics from both NS 1 and 2, but was distinctly different from 4.

Eight collaborating laboratories assayed 7 blind duplicate pairs of foods for polydextrose content. The 7 test sample pairs ranged from low (2%) to high (95%) levels. The following foods were prepared with polydextrose mixed into the other ingredients and then baked, cooked, or otherwise prepared: milk chocolate candy, iced tea, sugar cookie, grape jelly, soft jellied candy, and powdered drink mix. Collaborators received a polydextrose standard to develop a calibration curve. The method determined polydextrose by ion chromatography, after removal of interfering food components (high molecular weight solubles). Repeatability standard deviations (RSD_r) ranged from 3.93 to 9.04%; reproducibility standard deviations (RSD_R) ranged from 4.48 to 14.06%. The average recovery was 94%.