Molecular Characterization of Barley β-Glucans by Size-Exclusion Chromatography with Multiple-Angle Laser Light Scattering and Other Detectors

Molecular characteristics were determined for mixed-linkage (1→3) (1→4)-β-d -glucans (β-glucans) extracted from Azhul, Crystal, Waxbar, and Prowashonupana barleys. β-Glucans in extracts (with or without α-amylase, protease, hemicellulase, or xylanase treatment) were separated from other components by high-performance size-exclusion chromatography and detected with multiple-angle laser light scattering, refractive index, and fluorometry following postrefractive index treatment with Calcofluor. Pretreatment of barley with 70% ethanol (80°C, 4 hr) reduced β-glucanase activity by ~20%. Hot-alcohol treatment also reduced β-glucan extraction at 23 and 65°C by 42 and 14%, respectively. Molecular weights of β-glucans in the first water extract were generally higher than in succeeding water and alkali extracts. Weight average molecular weights ranged from 0.44 × 10⁶ to 2.34 × 10⁶ g/mol after α-amylase treatment to remove interfering starch. Interference due to pentosans was not demonstrated using enzyme treatments.     

Preparation and Solubility of Phosphorylated β-Cyclodextrins

β-Cyclodextrin (CD) was phosphorylated with phosphoryl chloride in aqueous alkaline media at different temperatures and pH values. The phosphorylated cyclodextrin (PCD) were characterized by phosphorus contents and positions of substitution as determined by ³¹P-NMR spectroscopy. Reaction of CD with equivmolar POCl₃ for 3 hr at pH 12 and 45°C yielded in a PCD with a phosphorus content of 5.67%. The ratio of monoand diphosphate esters increased when the reaction temperature was raised from 25 to 60°C. The monoesterified phosphate groups were mainly located at C-6 of the anhydroglucose units when the reaction pH was 11 or 12. Reactions at pH 10, however, led to a higher degree of substitution at C-2 than at C-6. Phosphorylation enhanced the water solubility of CD. Solubility of a PCD (5.65% phosphorus) was 35% at pH 8 and 25°C. Simultaneously, solubility of the PCD in 25% ethanol in water was much greater than unsubstituted CD (22.3 vs. 2.8%). The PCD enhanced the water solubility of nonpolar compounds, such as β-carotene.     

Molecular Characterization of Corn Starch Using an Aqueous HPSEC‐MALLS‐RI System Under Various Dissolution and Analytical Conditions

Molecular characteristics based on absolute weight‐average molecular weight (M_w) and z‐average radius of gyration (R_g) of normal corn starch were analyzed by high‐performance size‐exclusion chromatography (HPSEC) attached to multiangle laser‐light scattering (MALLS) and refractive index (RI) detectors under different starch dissolution and analytical conditions. Autoclaving (121°C, 20 min) or microwave heating (35 sec) provided better HPSEC recovery and higher M_w for starch molecules than simple dissolution in hot water. The M_w for the autoclaved corn amylopectin and amylose fractions separated with a TSK G5,000 column at 60°C were 201 × 10⁶ and 3.3 × 10⁶, respectively. The specific volume for gyration (SV_g) calculated from M_w and R_g could be used for the comparison of molecular compactness which was inversely related to the degree of branching. The SV_g values of amylopectin and amylose fractions in the chromatogram (TSK G5,000, autoclaved for 20 min) were 0.092 and 0.529, respectively. But a portion (20–30%) of large amylopectin molecules did not pass the injection membrane filter (3.0 μm) and the SEC column, resulting in incomplete recovery. The unfiltered portion varied according to the dissolution treatment. Homogenization (7,000 rpm, 5 or 10 min) of the starch solution improved the recovery of the amylopectin fraction, but significantly increased the M_w of the amylose fraction (17 × 10⁶). Sonication for 5 min degraded starch molecules. For accurate analysis of a native starch using an aqueous SEC, the starch should be fully dissolved with proper treatment such as autoclaving or microwaving, and the column should be improved for full recovery of large amylopectin molecules.     

Effect of Extraction Conditions on Yield,Composition, and Viscosity Stability of Barley β-Glucan Gum

Cereal β-glucan can function as a thickener, but endogenous β-glucanase enzymes of the grain cleave β-glucan, reducing its viscosity. Although different extraction techniques have been developed, the viscosity stability of β-glucan gum has not been reported. The objective of this study was to investigate the effect of extraction treatments on the yield, purity, and viscosity stability of barley β-glucan (BBG) gum. A regular barley cultivar, Condor, and a waxy cultivar blend were extracted at pH 7–10 and 55°C for 0.5 hr. Four extraction conditions were evaluated: 1) extraction at high pH with no additional heat treatment; 2) boiling of extract; 3) prior refluxing of flour with 70% ethanol; and 4) treatment of extract with thermostable α-amylase for purification. Viscosity of extracts was monitored for ≥24 hr at 25°C. The highest β-glucan purities were achieved with a boiled Condor extract at pH 7 (81.3% db, 4.1% yield) and with refluxed waxy barley extracted at pH 8 and treated with α-amylase and (79.3% db, 5.1% yield). Gums extracted without subsequent heat treatment or prior refluxing of flour had high protein (>17%) and starch (>24%) impurities, respectively. The viscosity of gums obtained without heating was unstable. Prior refluxing treatment was not sufficient to stabilize final extracts. Boiling extracts resulted in stable but low viscosity. Reflux followed by purification treatment produced the highest stable viscosity for 0.5% solutions of both Condor (64 mPa sec^-1, pH 7) and waxy (48.8 mPa sec^-1, pH 8) extracts. Stable BBG gum with high viscosity can be obtained using thermal treatments in combination with high pH. The potential use of such gums as thickeners in food systems needs to be assessed.     

Large-Scale Purification and Characterization of Barley Limit Dextrinase,a Member of the α-Amylase Structural Family

Homogeneous barley limit dextrinase (LD) was isolated on a large scale in a yield of 9 mg/kg of 10-day germinated green malt. This represents a 9,400-fold purification and 29% recovery of the activity in a flour extract in 0.2M NaOAc (pH 5.0) containing 5 mM ascorbic acid. The purification protocol consists of precipitation from the extract at 20–70% saturated ammonium sulfate (AMS), followed by diethylaminoethyl (DEAE) 650S Fractogel anion-exchange chromatography, and affinity chromatography on β-cyclodextrin-Sepharose in the presence of 2M AMS. LD was eluted by 7 mMβ-cyclodextrin and contains a single polypeptide chain of 105 kDa (SDS-PAGE) and pI 4.3. Sequence analysis of tryptic fragments, prepared from 2-vinylpyridinylated LD and purified by RP-HPLC, identified short motifs recognized in β-strand 2, 3, and 5 characteristic of a catalytic (β/α)₈-barrel domain of the α-amylase family of amylolytic enzymes. Barley LD has ≈50 and 85% sequence identity to bacterial pullulanases and rice starch debranching enzyme, respectively. By using ¹H-NMR spectroscopy, LD hydrolyzes specifically α-1,6-glucosidic linkages in pullulan and a branched oligodextrin, 6²-O-α-maltotriosyl-maltotriose, with retention of the α-anomeric configuration. β-Cyclodextrin competitively inhibits the LD activity with K_i of 40 μM, while K_i is 1.9 mM and 2.4 mM for α-cyclodextrin and γ-cyclodextrin, respectively.     

Development of a Germination Process for Producing High β‐Glucan,Whole Grain Food Ingredients from Oat

Germination can be used to improve the texture and flavor of cereals. However, germination generally causes breakdown of β‐glucans, which is undesirable with respect to the functional properties of β‐glucan. Our aim was to assess possibilities of germinating oat without substantial loss of high molecular weight β‐glucan. Two cultivars, hulled Veli and hull‐less (naked) Lisbeth were germinated at 5, 15, and 25°C and dried by lyophilization or oven drying. Elevated germination temperatures led to an increase in Fusarium, aerobic heterotrophic bacteria, Pseudomonas spp., lactic acid bacteria, enterobacteria, and aerobic spore‐forming bacteria. Therefore, the germination temperature should be kept low to avoid excessive growth of microbes. Of the samples germinated at 15°C, only one contained low amounts of the Fusarium toxin deoxynivalenol (52 μg/kg). Germination led to the breakdown of β‐glucans, but the decrease in the molecular weight of β‐glucan was initially very slow. A short germination schedule (72 hr, 15°C) terminated with oven drying was developed to produce germinated oat with retained β‐glucan content. Compared with the native oat, 55–60% of the β‐glucan could be retained.     

Molecular Weight Distribution and (1→3)(1→4)-β-d-Glucan Content of Consecutive Extracts of Various Oat and Barley Cultivars

The content and molecular weight (MW) of β-glucan in extracts from a selection of oat and barley cultivars were compared using flow-injection analysis and high-performance size-exclusion chromatography. From 60 to 75% of the β-glucan was extracted from oat and waxy barley by hot water (90°C) containing heat-stable α-amylase, whereas just 50–55% was extracted from nonwaxy barley. Consecutive extractions with hot water and dimethylsulfoxide (DMSO) extracted 65% (nonwaxy barley) or 75–80% (oat and waxy barley) of the total β-glucan. An extraction with sodium hydroxide and sodium borohydride (NaOH/NaBH₄) increased the percentage of β-glucan extracted to 86–100% but decreased the MW. The MW of β-glucan in the oat cultivars selected was significantly higher than those in the barley cultivars. The β-glucan extracted from the nonwaxy barley cultivars showed significantly higher peak MW than that from the waxy barley cultivars.     

Effect of Partial Acid Hydrolysis and Heat‐Moisture Treatment on Formation of Resistant Tuber Starch

Structural characteristics of resistant starch (RS) were investigated. Tuber starches, hydrolyzed with 1N HCl at 35°C for 8 hr followed by autoclaving‐cooling treatment, were heated at 100°C for 16 hr after adjusting the moisture content to 20 or 30%. RS content of the tuber starches ranged from 5.4 to 22.7% depending on the source and type of treatment. Gelatinization parameters of RS isolated from partially acid‐hydrolyzed starch with autoclaving‐cooling followed by heat‐moisture treatment (HMT) showed higher enthalpy (ΔH) values and lower peak temperature (T_p) compared with non‐acid‐hydrolyzed RS. R values, the difference between completion and initial temperatures, and ΔH of RS increased by HMT. The X‐ray diffraction patterns of potato and sweet potato RS isolated from partially acid‐hydrolyzed starch with autoclaving‐cooling showed distinct sharp peaks at 15, 25, 27, and 28°, which decreased by HMT.     

Light-Scattering Molecular Weights and Intrinsic Viscosities of Processed Waxy Maize Starches in 90% Dimethyl Sulfoxide and H2O

Waxy maize starch was treated by a variety of gentle and severe methods: direct dispersion-solubilization into 90% dimethyl sulfoxide (DMSO) and H₂O solvent, extrusion followed by dispersion-solubilization of the ground exudate into solvent, or jet-cooking or stirred autoclaving of an aqueous starch slurry followed by transfer into solvent. Intrinsic viscosities [η] and multiangle light-scattering measurements were made in 90% DMSO-H₂O. A Mark-Houwink relation, [η] = (0.28–1.2) M_w^{0.29 ± 0.04}, was obtained over a molecular weight range of ≈30–700 million. However, there was a large amount of scatter in the data when [η] were >140 mL/g. The power law relationship R_g ∝ M_w^0.41±0.04 was noted between radii of gyration and molecular weights. We infer from our data that over the entire range of M_w distributions, the amylopectin existed in solution as relatively compact molecules or aggregates and that in the higher molecular weight region, the size and possibly the shape of the “dissolved” amylopectin was highly sensitive to the method of dispersal and treatment.     

Rheological Properties of (1→3),(1→4)-β-d-Glucans from Raw,Roasted, and Steamed Oat Groats

Effects of hydrothermal treatments (steaming, roasting) of oat grain on β-glucan extractability and rheological properties were tested on oat cultivars with low (Robert) and high (Marion) β-glucan content. Steaming of grain reduced the amount of β-glucan that could be extracted, compared with raw or roasted grain, but the extracts from steamed grain had much greater viscosity. Increased extraction temperatures increased the amount and the average relative molecular mass (M _r) value of β-glucan extracted. In boiling water extractions, the average M _r values among raw, roasted and steamed oat samples were equivalent, but extracts from steamed oat grain had significantly higher intrinsic viscosity than the extracts from roasted or raw oat grains. β-glucan solutions purified from steamed grain extracts were very viscous and highly pseudoplastic, as described by the power law equation. Oat β-glucan from steamed samples were more viscoelastic than β-glucan from roasted or raw oat samples. Because viscous properties of β-glucan from boiling water extracts are influencedhydrothermal treatments without affecting polymer molecular weight, polymer interaction with the solvent must be affected. Steaming may disrupt intramolecular cross-linkings in native β-glucan, allowing a linear chain configuration to generate greater viscosity.     

Synergistic Hydrolysis of Crude Corn Starch by α‐Amylases and Glucoamylases of Various Origins

Four α‐amylases and two glucoamylases from various sources, in eight combinations, were used to study the synergistic hydrolysis of crude corn starch at various temperatures. At 40 and 50°C, the combinations containing Rhizopus mold glucoamylase enhanced hydrolysis of corn starch compared wth that obtained with the combinations from Aspergillus niger. At 60°C, Rhizopus mold combinations gave low reaction yields as the enzyme was inactivated. The differences observed between α‐amylases are smaller, with the exception of Bacillus licheniformis α‐amylase, which presented more than twice the productivity of the other α‐amylases, at all temperatures. In terms of substrate conversion at 5 hr of hydrolysis, the combination of B. licheniformis α‐amylase with Rhizopus mold glucoamylase at 50°C presents 76% substrate conversion, whereas, with all the other combinations, starch conversion was 13–73%. HPLC analysis of the reaction products obtained at 50°C showed that the main product of corn starch hydrolysis was glucose at 85–100%. Further experiments showed that A. niger glucoamylase and B. licheniformis α‐amylase were the only enzymes that retained their initial activity after incubation at the temperatures studied.     

Molecular Characterization of Wheat Amylopectins by Multiangle Laser Light Scattering Analysis

Weight-average molecular weight (M_w) and chain length of eight amylopectins isolated from one Australian, two United States, and five Korean wheats were measured using multiangle laser light scattering (MALLS) and refractive index (RI) detectors operated in a microbatch mode, and in a high-performance size-exclusion chromatography (HPSEC) mode. The M_w of amylopectins measured in the microbatch mode ranged from 29 × 10⁶ to 349 × 10⁶. Three amylopectins (Geuru, Tapdong and WW) showed significantly high M_w values over 200 × 10⁶. The M_w measured by HPSEC mode with MALLS-RI detectors (42 × 10⁶ to 73 × 10⁶) were significantly less than those obtained in the microbatch mode with exception of dark northern spring hard wheat (DNS) amylopectin, indicating the possible variation of M_w by the analysis mode. Root-mean square of the radius of gyration (R_g) also was greater when the microbatch mode was used (122–340 vs. 95–116 nm). Chain length distributions of debranched amylopectins of different cultivars, measured by the HPSEC-MALLS-RI system, were similar. Weight average degrees of polymerization (DPw) of A, B₁, and larger B chains (B_≥2) had ranges of 13–22, 26–46, and 58–73, respectively, and mass ratios of A and B chains ranged from 0.7 to 1.1.     

Hot‐Water Solubilities and Water Sorptions of Resistant Starches at 25°C

Resistant starches (RS) were prepared from wheat starch and lintnerized wheat starch by autoclaving and cooling and by cross‐linking. Heat‐moisture treatment also was used on one sample to increase RS. The experimental resistant starches made from wheat starch contained 10–73% RS measured as Prosky dietary fiber, whereas two commercial resistant starches, Novelose 240 and 330, produced from high‐amylose maize starch, contained 58 and 40%, respectively. At 25°C in excess water, the experimental RS starches, except for the cross‐linked wheat starch, gained 3–6 times more water than the commercial RS starches, and at 95°C gained 2–4 times more. Cross‐linked RS4 wheat starch and Novelose 240 showed 95°C swelling powers and solubilities of 2 g/g and 1%, and 3 g/g and 2%, respectively. All starches showed similar water vapor sorption and desorption isotherms at 25°C and water activities (a_w) < 0.8. At a_w 0.84–0.97, the resistant starches made from wheat starch, except the cross‐linked wheat starch, showed ≈10% higher water sorption than the commercial resistant starches.     

Physical and Sensory Characteristics of Extruded Products Made from Two Oat Lines with Different β‐Glucan Concentrations

The impact of extrusion on physical and sensory properties and on the in vitro bile acid (BA) binding was examined for N979 and Jim oat (Avena sativa) lines with 8.1 and 4.8% β‐glucan, respectively. Based on hardness and edibility of products made from Jim oats, moisture concentrations of 16–25% and temperatures of 165–180°C were selected for N979 extrusion. Jim‐based cereal had a significantly greater (P < 0.05) expansion ratio than did N979‐based cereal at most moistures. N979 cereal was browner, but not harder, than Jim cereal. Extruded products from N979 and Jim oats had 5.29–5.99% and 3.38–3.94% β‐glucan, respectively. Changing extrusion temperature or moisture content did not affect β‐glucan concentration in the products. N979 cereal made at 165°C and 16% moisture had greater BA binding than at other conditions, and had crunchiness comparable to cereals made at other conditions. BA binding of Cheerios brand breakfast cereal was close to that of N979 cereal made at 180°C and 18% moisture, but lower than cereals made at other conditions. Cereals made from Jim and N979 oats were browner, harder, coarser, and crunchier than Cheerios breakfast cereal. Proper processing and preparation techniques should be considered when producing extruded products from high β‐glucan oats.     

Glycemic Response to Oat Bran Muffins Treated to Vary Molecular Weight of β‐Glucan

Oat bran muffins, containing 4 or 8 g of β‐glucan per two‐muffin serving, were prepared with or without β‐glucanase treatment to produce a range of β‐glucan molecular weights from 130,000 to just over 2 million. Following an overnight fast, the glycemic responses elicited by the untreated and treated muffins was measured in 10 healthy subjects and compared with a control whole wheat muffin. Taken all together, the 4‐g β‐glucan/serving muffins reduced blood glucose peak rise (PBGR) by 15 ± 6% compared with the control. The 8‐g β‐glucan/serving muffins had a significantly greater effect (44 ± 5% reduction compared with the control, P < 0.05). The efficacy of the muffins decreased as the molecular weight was reduced from a 45 ± 6% reduction in PBGR (P < 0.05) for the untreated muffins (averaged of both serving sizes) to 15 ± 6% (P < 0.05) for muffins with the lowest molecular weight. As the molecular weight was reduced from 2,200,000 to 400,000, the solubility of the β‐glucan increased from a mean of 44 to 57%, but as the molecular weight was further decreased to 120,000, solubility fell to 26%. There was a significant correlation (r² = 0.729, P < 0.001) between the peak blood glucose and the product of the extractable β‐glucan content and the molecular weight of the β‐glucan extracted.     

Cross‐Linked Resistant Starch: Preparation and Properties

Resistant starches (RS) were prepared by phosphorylation of wheat, waxy wheat, corn, waxy corn, high‐amylose corn, oat, rice, tapioca, mung bean, banana, and potato starches in aqueous slurry (≈33% starch solids, w/w) with 1–19% (starch basis) of a 99:1 (w/w) mixture of sodium trimetaphosphate (STMP) and sodium tripolyphosphate (STPP) at pH 10.5–12.3 and 25–70°C for 0.5–24 hr with sodium sulfate or sodium chloride at 0–20% (starch basis). The RS₄ products contain ≤100% dietary fiber when assayed with the total dietary fiber method of the Association of Official Analytical Chemists (AOAC). In vitro digestion of four RS₄ wheat starches showed they contained 13–22% slowly digestible starch (SDS) and 36–66% RS. However after gelatinization, RS levels fell by 7–25% of ungelatinized levels, while SDS levels remained nearly the same. The cross‐linked RS₄ starches were distinguished from native starches by elevated phosphorus levels, low swelling powers (≈3g/g) at 95°C, insolubilities (<1%) in 1M potassium hydroxide or 95% dimethyl sulfoxide, and increased temperatures and decreased enthalpies of gelatinization measured by differential scanning calorimetry.     

Structural and Functional Characteristics of Selected Soft Wheat Starches

Starches from eight soft wheat samples (two parent lines and six offspring) were isolated; relationships between their structures and properties were examined. Branch chain‐length distributions of amylopectins were determined by using high‐performance anion exchange chromatography equipped with an amyloglucosidase reactor and a pulsed amperometric detector (HPAEC‐ENZ‐PAD). Results showed that the average chain length of the eight samples varied at DP 25.6–26.9. Starch samples of lines 02, 60, 63, 95, and 114 consisted of amylopectins with more long chains (DP ≥ 37) and longer average chain length (DP 26.2–26.9) than that of other samples. These starch samples of longer branch chain length displayed higher gelatinization temperatures (55.3–56.5°C) than that of other samples (54.4–54.9°C) and higher peak viscosity (110–131 RVU) and lower pasting temperature (86.3–87.6°C) than others (83–100 RVU and 88.2–88.9°C, respectively). The M_w of amylopectins, determined by using high‐performance size exclusion chromatography equipped with multiangle laser‐light scattering and refractive index detectors (HPSEC‐MALLS‐RI), were similar for all samples (6.17 × 10⁸ to 6.97 × 10⁸). There were no significant differences in amylose and phosphorus contents between samples. These results indicated that physical properties of wheat starch were affected by the branch‐chain length of amylopectin.     

Separating cellular metabolism from exoenzyme activity in soil organic matter decomposition

Soil organic matter (SOM) decomposes both inside and outside of cells. Cellular metabolism and extracellular depolymerization normally operate simultaneously in soil but are difficult to separate in practice. To learn more about the extracellular component of SOM decomposition, we sterilized a semiarid annual grassland soil to inhibit cellular metabolism, and then assayed cell viability, exoenzyme activities, and pathways of carbon dioxide (CO₂) emission. Chloroform (CHCl₃) fumigation was intended to disrupt cellular activities while leaving biochemical processes intact. Gamma (γ) irradiation and autoclaving were intended to disrupt both cellular and extracellular biochemical processes while leaving abiotic processes intact. We measured the potential activities of eight enzymes (six hydrolytic, two oxidative) and CO₂ emission induced by seven substrates (glucose, three amino acids, three tricarboxylic acid [TCA] cycle intermediates). We found that all three sterilization techniques clearly disrupted cellular metabolism. Chloroform and irradiation decreased cultivable cell counts by 2–3 orders of magnitude, inhibited CO₂ emission pathways associated with glucose and amino acids, and decreased the hydrolytic activities of α-glucosidase and xylosidase by 72–82%. The other hydrolytic enzymes (β-glucosidase, cellobiohydrolase, NAGase, phosphatase) were less sensitive to both CHCl₃ and irradiation. All hydrolytic activities that we assayed were inhibited by autoclaving, indicating that biochemical reactions and other extracellular processes drive hydrolytic SOM decomposition. Oxidative activities, on the other hand, did not stop after autoclaving or even combusting at 500 °C. This supports other studies which have found that mineral catalysts partly drive oxidative SOM decomposition. Unexpectedly, CO₂ emission from TCA intermediates decreased by only 26–47% after sterilization suggesting that the required dehydrogenase enzymes for decarboxylation are still active when cells are dead but relatively intact. Because CHCl₃ had slightly smaller effects on exoenzyme activities compared to irradiation, and because it may be continuously applied, limiting the potential for recolonization and regrowth (unlike irradiation), we suggest it is an adequate and more accessible method for separating the activity of exoenzymes from cellular metabolism under realistic soil conditions.     

Analysis of 2‐Acetyl‐1‐Pyrroline in Rice by HSSE/GC/MS

An extremely sensitive method for the analysis of 2‐acetyl‐1‐pyrroline (2AP) in rice, employing stir bar sorptive extraction (Twister) was studied. The Twister stir bar is placed in the headspace of a 20‐mL vial containing 1 g of rice kernels, 7.5 mL of 0.1M KOH, and 2.2 g of NaCl, along with a second Teflon‐coated stir bar for mixing. Analytes are adsorbed onto the Twister for 4 hr at 40°C and then desorbed at 270°C into a GC column while cryofocusing at –80°C. The headspace sorptive extraction (HSSE) method was able to detect <0.1 ppb of 2AP in rice. The precision of the HSSE method (>10%) was not as good as the GC/FID method (≈6%). Using HSSE, 2AP was observed in all samples generally considered to be aromatic and was not observed in any nonaromatic samples. Additionally, a modified method for the synthesis of 2‐acetyl‐1‐pyrroline was studied and the presence of a tautomer of 2‐acetyl‐1‐pyrroline was confirmed.     

Retrogradation Mechanism of Rice Starch

The non‐Newtonian behavior and dynamic viscoelasticity of rice starch (Akihikari, 18.8% amylose content) solutions after storage at 25 and 4°C for 24 hr were measured with a rheogoniometer. The flow curves, at 25°C, of Akihikari starch showed plastic behavior >3.0% (w/v) after heating at 100°C for 30 min. The dynamic viscoelasticity of the starch increased after storage at 25 and 4°C for 24 hr and stayed at a constant value with increasing temperature. A small dynamic modulus of rice starch was observed upon addition of urea (4.0M) at low temperature (0°C), but it produced a sigmoid curve when plotted against increasing temperature. A small dynamic modulus was also observed in 0.05M NaOH solution. However, it increased rapidly after the temperature reached 70°C. Possible models of retrogradation mechanism of rice starch were proposed.