Preparation and characterization of new and improved soluble-starches, -amylose, and -amylopectin by reaction with benzaldehyde/zinc chloride

Seven different starches from potato, rice, maize, waxymaize, amylomaize-VII, shoti, and tapioca, and potato amylose and potato amylopectin have been reacted with benzaldehyde, catalyzed by ZnCl₂, to give new water-soluble starches and water soluble-amylose and soluble-amylopectin. In contrast to the native starches, aqueous solutions of the modified starches could not be precipitated with 2-, 3-, or 4-volumes of ethanol. β-Amylase gave no reaction with the modified starches, in contrast to the native starches, indicating that the modification occurred exclusively at the nonreducing-ends, giving 4,6-benzylidene-d-glucopyranose at the nonreducing-ends. Reactions of α-amylase with native and modified potato and rice starches gave a decrease in the triiodide blue color and an increase in the reducing-value that were similar for the native- and modified-starches, indicating the modified starches had not been significantly altered by the modification. The benzaldehyde-modified starches and benzaldehyde-modified potato amylose and potato amylopectin components, therefore, have a starch structure very much like their native counterparts, in contrast to the Lintner, Small, and the alcohol/acid-hydrolyzed soluble-starches that have undergone acid hydrolysis. The benzaldehyde-modified starches and starch components have significantly higher water solubility than their native counterparts even though the structures of the modified starches had only been slightly altered from the structures of their native counterparts. They all gave crystal-clear solutions that did not retrograde.     

Structural and physicochemical properties of starch gels prepared from partially modified starches using Thermus aquaticus 4-α-glucanotransferase

The modified starch gels prepared from partial enzyme treatments (1, 3, and 6 U/g starch; 2-h incubation) of the corn and rice starch pastes using Thermus aquaticus 4-α-glucanotransferase (TAαGT) were investigated for their molecular characteristics, microstructures, and physicochemical properties. Unlike the native and partially modified normal starches, the native and partially modified waxy starches could not form gels strong enough for textural analysis after 24 h for gel setting. Features of the partially modified normal starches were the specific apparent amylose contents and maximum iodine absorption wavelength (λ_max, ∼567 nm), as well as the tri-modal molecular weight profiles and flatter side-chain distributions. Also, the partially modified normal starch gels possessed fractured surfaces with discontinuous crystalline fibrous assembly that differed from the native starch gels’ porous continuous network, which resulted in more brittle, rigid, and resilient gels compared with the native gels.     

Structural characteristics and physicochemical properties of oxidized corn starches varying in amylose content

The effects of amylose content on the extent of oxidation and the distribution of carboxyl groups in hypochlorite-oxidized corn starches were investigated. Corn starches including waxy corn starch (WC), common corn starch (CC), and 50% and 70% high-amylose corn starches (AMC) were oxidized with NaOCl at three concentrations (0.8%, 2%, and 5%). Carboxyl and carbonyl content of oxidized starches increased with increasing NaOCl concentration. High-AMC (70%) had slightly higher carboxyl and carbonyl contents at 0.8% NaOCl, whereas WC had significantly higher carboxyl and carbonyl contents at 2% and 5% NaOCl levels. Carbohydrate profiles by high-performance size-exclusion chromatography indicate that amylose was more susceptible to depolymerization than amylopectin. Degradation of amylopectin long chains (DP >24) was more pronounced in WC and CC than in AMCs. The crystalline lamellae of WC started to degrade at 2% NaOCl, but those of the other corn starches remained intact even at 5% NaOCl level according to X-ray crystallinity. By using anion-exchange chromatography for separation and size-exclusion chromatography for characterization, carboxyl groups were found to be more concentrated on amylopectin than on amylose, particularly in AMCs. Oxidation decreased gelatinization temperature and enthalpy with WC showing the most decrease and 70% AMC showing the least. The gelatinization enthalpy of 50% AMC decreased significantly faster than those of CC and 70% AMC after 0.8% oxidation. Retrogradation of amylopectin slightly increased after oxidation with increasing oxidation level. The peak viscosities of oxidized WC and CC were higher than those of their native counterparts at 0.8% NaOCl, but this increase was not observed in AMCs. The setback viscosities of 2% NaOCl-oxidized 50% and 70% AMCs were much higher than those of the unmodified counterparts. The extent of oxidation and physicochemical properties of oxidized starches varied greatly with the amylase:amylopectin ratio of corn starches. Amylose was suggested to play an important role in controlling the oxidation efficiency.     

Preparation and characterization of soluble starches having different molecular sizes and composition,by acid hydrolysis in different alcohols

Potato and waxy-maize starches were separately modified for 1 h at 65° with 0.36% hydrochloric acid in methanol, ethanol, 2-propanol, and 1-butanol. All of the modified starches were readily soluble in hot water, to give crystal-clear solutions up to a concentration of at least 20% (w/v). The modified granules were studied by light-microscopy and iodine-iodide staining. All of the modified starches retained their granule appearance, although with various degrees of damage that progressively increased from methanol to 1-butanol. Both hydrolysis and alcoholysis occurred, but to different extents in the different alcohols. The highest proportion of alcoholysis occurred in methanol where 50% of the resulting molecules were glycosides, the lowest in 1-butanol where 6% were glycosides. The number-average molecular weights of the modified starches also progressively decreased from 126,670 for the methanol-modified waxy-maize starch to 4,750 for the 1-butanol-modified potato starch. The methanol- and ethanol-modified potato starches were fractionated into amylose and amylopectin components. The 2-propanol- and 1-butanol-modified potato starches gave only an amylopectin component. The amylose components were characterized by gel-permeation chromatography on Bio-Gel A-5m, and the amylopectin components, on Bio-Gels A-150m and A-0.5m. The molecular sizes of the amylose and amylopectin components progressively decreased from methanol- to 1-butanol-modified starches. Furthermore, the polymodal composition of the amylopectin component was decreased to give a more homogeneous product. Waxy-maize starch was modified in methanol and 2-propanol and gave products that were of lower molecular size and more homogeneous than the polymodal native starch. It is shown that the differential effect of the different alcohols on the modification of the starch granules is produced by effecting different concentrations of acid inside the granule, where hydrolysis occurs in the 10–12% of water contained in the granule. It is postulated that 2-propanol and 1-butanol dissolve the double-helical, crystalline regions in the starch granule to give different types of products under otherwise identical conditions of modification.     

Charakterisierung der Stärkehydrolysate nach Einwirkung einer thermostabilen α-Amylase

The action of thermostable α-amylase produced by Bacillus licheniformis 44MB82 strain on soluble and insoluble starch, amylose and amylopectin at temperatures 30°C and 90°C was studied. The hydrolysis of soluble starch proceeded rapidly for 10 to 15 minutes after which the maltodextrins thus formed were further dissociated. In the course of 60-minutes enzyme treatment mainly glucose, maltose and maltosugars (from G₃ to G₆) as low molecular weight products were found and the formation of maltcse and maltotriose was increased by the longer treatment. The hydrolysis of insoluble starch and amylopectin proceeded in the same way while the amylose was hydrolysed slowly.     

Pasting viscosity and in vitro digestibility of retrograded waxy and normal corn starch powders

Pasting viscosity and in vitro digestibility of oven-dried powders of waxy and normal corn starch gels (40% solids) retrograded under an isothermal (4 °C) or temperature cycled (4/30 °C) storage were investigated. Temperature cycling induced higher onset temperature for melting of amylopectin crystals than isothermal storage under a differential scanning calorimeter whereas little difference in crystalline type was observed under X-ray diffraction analysis. Temperature cycling caused higher pasting temperature and viscosity for the retrograded starches than isothermal storage. The retrograded waxy corn starch powders exhibited pasting behaviors similar to that of native waxy corn starch. However, the retrograded normal corn starch powders showed very much different pasting patterns with lower pasting viscosity but higher pasting temperature than native starch counterpart. The retrogradation increased slowly digestible starch content without changing resistant starch content, more effectively by the temperature cycling than the isothermal storage.     

Locations of hypochlorite oxidation in corn starches varying in amylose content

The general oxidation mechanism by hypochlorite on starch has been well studied, but the information on the distribution of the oxidation sites within starch granules is limited. This study investigated the locations where the oxidation occurred within corn starch granules varying in amylose content, including waxy corn starch (WC), common corn starch (CC), and 50% and 70% high-amylose corn starch (AMC). Oxidized corn starches were surface gelatinized by 13 M LiCl at room temperature to different extents (approximately 10%, 20%, 30%, and 40%). The surface-gelatinized remaining granules were separated and studied for structural characteristics including carboxyl content, amylose content, amylopectin chain-length distribution, thermal properties, and swelling properties. Oxidation occurred mostly at the amorphous lamellae. More carboxyl groups were found at the periphery than at the core of starch granules, which was more pronounced in oxidized 70% AMC. More amylose depolymerization from oxidation occurred at the periphery of CC. For WC and CC, amylopectin long chains (>DP 36) were more prone to depolymerization by oxidation. The gelatinization properties as measured by differential scanning calorimetry also supported the changes in amylopectin fine structure from oxidation. Oxidized starches swelled to a greater extent than their unmodified counterparts at all levels of surface removal. This study demonstrates that the locations of oxidation and physicochemical properties of oxidized starches are affected by the molecular arrangement within starch granules.     

Starch hydrolysis modeling: application to fuel ethanol production

Efficiency of the starch hydrolysis in the dry grind corn process is a determining factor for overall conversion of starch to ethanol. A model, based on a molecular approach, was developed to simulate structure and hydrolysis of starch. Starch structure was modeled based on a cluster model of amylopectin. Enzymatic hydrolysis of amylose and amylopectin was modeled using a Monte Carlo simulation method. The model included the effects of process variables such as temperature, pH, enzyme activity and enzyme dose. Pure starches from wet milled waxy and high-amylose corn hybrids and ground yellow dent corn were hydrolyzed to validate the model. Standard deviations in the model predictions for glucose concentration and DE values after saccharification were less than ±0.15% (w/v) and ±0.35%, respectively. Correlation coefficients for model predictions and experimental values were 0.60 and 0.91 for liquefaction and 0.84 and 0.71 for saccharification of amylose and amylopectin, respectively. Model predictions for glucose (R ² = 0.69–0.79) and DP4⁺ (R ² = 0.8–0.68) were more accurate than the maltotriose and maltose for hydrolysis of high-amylose and waxy corn starch. For yellow dent corn, simulation predictions for glucose were accurate (R ² > 0.73) indicating that the model can be used to predict the glucose concentrations during starch hydrolysis.     

Determination of the maximum water solubility of eight native starches and the solubility of their acidic-methanol and -ethanol modified analogues

The maximum water solubilities of eight native starches from potato, shoti, tapioca, maize, waxy maize, amylomaize-7, wheat, and rice and their acid-methanol and acid-ethanol modified analogues have been determined. Maximum solubilities of 18.7 and 17.4 mg/mL were obtained for waxy maize and tapioca and 12.4 mg/mL for potato and maize starches by autoclaving 220 mg/10 mL at 121 degrees C; 8.7 mg/mL was obtained for shoti starch by stirring in 85:15 (v/v) Me(2)SO-H(2)O at 20 degrees C; and 7.0 and 5.2mg/mL for rice and amylomaize-7 starches by stirring in 1M NaOH at 20 degrees C. The acid-alcohol treated starches were 4-9 times more soluble than their native starches. The compositions of the solubilized starches had, in general, much higher ratios of amylose to amylopectin than the ratios in their native granules. A major exception to this was the acid-methanol treated potato, shoti, and rice starches that had much lower ratios of amylose to amylopectin than the ratios in their granules.     

Sorption of Monoterpenoids and Sesquiterpenoids by Natural Polysaccharides

Sorption of terpenoids (essential oil components) from aqueous solutions by six types of native food starches was studied by capillary gas chromatography. Sorption of volatile substances did not depend on amylose content in starch and specific surface of its granules. The degree of sorption was maximum (86%) for corn starch containing 25–28% amylose and decreased in the following order: tapioca starch (77%) > potato starch (74%) > wheat starch (70%) > high-amylose corn starch (58%) > amylopectin corn starch (57%). Amylopectin corn starch differed from other starches in the mechanism of sorption and selectivity to compounds with various functional groups.     

Sorption of mono- and sesquiterpenes by natural polysaccharides

Sorption of terpenoids (essential oil components) from aqueous solutions by six types of native food starches was studied by capillary gas chromatography. Sorption of volatile substances did not depend on amylose content in starch and specific surface of its granules. The degree of sorption was maximum (86%) for corn starch containing 25-28% amylose and decreased in the following order: tapioca starch (77%) > potato starch (74%) > wheat starch (70%) > high-amylose corn starch (58%) > amylopectin corn starch (57%). Amylopectin corn starch differed from other starches in the mechanism of sorption and selectivity to compounds with various functional groups.     

Effect of sulfate and citrate salts on derivatization of amylose and amylopectin during hydroxypropylation of corn starch

Common corn starch was modified in 0.56 M sodium sulfate solution and in 0.31 M potassium citrate solution. It was found that about 1.8 times the amount of reagent (propylene oxide) was needed to get a same molar substitution (MS) when potassium citrate was used. Hydroxypropylated starches were fractionated on a size-exclusion column to separate amylose from amylopectin, and MS values of the whole starch, the amylose, and the amylopectin were determined. In all preparations, amylose was derivatized to a greater extent than was amylopectin. The data indicate that, with common corn starch: (1) the greater the overall derivatization, the greater was the preference for derivatization of amylose; and (2) the preference for amylose derivatization was greater for corn starch modified in potassium citrate solution than in sodium sulfate solution when the MS values for the two preparations were essentially the same.     

Starch phosphates prepared by reactive extrusion as a sustained release agent

Characteristics of native starch have limited its application in solid dosage forms as a sustained release agent. There is a growing interest in improving starch functionality for sustained release applications because of its non-toxicity and biodegradability. This study attempted to investigate extruded starch phosphates as an excipient in sustaining drug release. Starches from various botanical sources with different amylose contents, including waxy corn, common corn, Hylon V (50% amylose), Hylon VII (70% amylose), and potato, were used to prepare starch phosphates at pH 9.0 or 11.0 using a reactive extrusion method. Phosphorous content was higher for starch phosphates prepared at pH 9.0 than at pH 11.0, and varied with starch type when phosphorylated at pH 9.0. Reactive extrusion produced starch extrudates that upon forming hydrogels were capable of sustaining release of metoprolol tartrate (MPT). The structural features of the hydrogel as modified by the phosphorylation reaction were found to alter the kinetics of drug release from the swellable matrices. The unmodified extrudates formed weaker gels as evidenced by their rheological properties, and showed faster drug release. Waxy corn starch phosphorylated at pH 9.0 as well as common corn and potato starches phosphorylated at pH 11.0 were found to exhibit more case-II-like properties attributed to a high density of cross-links and stronger chain entanglement. Waxy corn starch phosphorylated at pH 9.0 exhibited the lowest degree of drug release. The entanglement among amylopectin molecules and branch chains was suggested to play a role in governing MPT release.     

Dynamic and extensional properties of starch in aqueous dimethylsulfoxide

The effect of starch composition and concentration on the rheological properties of starch in a mixed solvent, water–DMSO, was investigated in dynamic shear and extensional mode. High amylose corn starch containing 70% amylose and 30% amylopectin, common corn starch containing 25% amylose and 75% amylopectin, and waxy corn starch containing about 99% amylopectin were used in this study. Concentrations of 2, 4, 6, and 8% (w/v) in 10% water-90% DMSO (v/v) were used for each starch type. An increase in the amylopectin content of starch from 30 to 99% caused a change in behavior from semidilute solution to viscoelastic solid at a concentration of 8% (w/v). At a concentration of 2%, an increase in the amylopectin content of starch from 30 to 99% caused a change from Newtonian to incipient gel-like behavior. Behavior at intermediate concentrations of 4 and 6% (w/v) varied from semidilute to critical gel-like with increasing amylopectin content. A power-law relaxation was observed for all concentrations of common and waxy corn starches with the slope decreasing with increase in concentrations. A 2% waxy corn starch solution displayed extension thinning behavior, while a 2% high amylose corn starch solution displayed Newtonian behavior.     

Purification and characterization of β-amylase from Curculigo pilosa

Curculigo pilosa is traditionally used in the manufacture of sorghum beer in West Africa. β-Amylase was purified 100-fold with 38% yield from a crude extract, giving final specific activities of 4850 U/mg and 5650 U/mg using soluble starch and p-nitrophenyl maltopentaoside, respectively, as substrates. The molecular mass of the monomeric enzyme was 64 kDa and its pI 4.2. Both activity and thermostability are higher than reported for other plant β-amylases. The catalytic efficiency was lower for amylose than for starches and amylopectin. In contrast to other plant amylases, the β-amylase from C. pilosa is able to degrade raw starches from wheat, corn, potato and rice. In this respect, it resembles β-amylases from microbial origin. This property, and its high activity and stability, explain its traditional use in the manufacture of infant food and sorghum beer in Burkina Faso and could make it applicable for other biotechnological purposes. Received: 4 March 1999 / Received revision: 6 August 1999 / Accepted: 13 August 1999     

Effect of different substrates and their preliminary treatment on cyclodextrin production

Summary Various kinds of substrates were tested for cyclodextrin production with cyclodextrin glucanotransferase (CGTase) from Bacillus megaterium. The enzyme formed cyclodextrin from different kinds of starch, dextrins, amylose, and amylopectin. However, the highest degree of conversion was obtained from starch. Corn starch appeared to be the best substrate – the cyclodextrin yield was 50.9%. The effect of molecular mass and preliminary treatment of starch with α-amylase on the CD yield was investigated. It was proved that CGTase preferred native starch with high molecular mass and low dextrose equivalent. The preliminary treatment with α-amylase occurred to be inefficient and unnecessary since it did not lead to an increase in the CD yield. Some of the substrates were treated with pullulanase. The effect of debranching was highest in the case of corn starch: the cyclodextrin yield increased by 10%.     

Internal structure and physicochemical properties of corn starches as revealed by chemical surface gelatinization

The organization of amylose and amylopectin within starch granules is still not well elucidated. This study investigates the radial distribution of amylose and amylopectin in different corn starches varying in amylose content (waxy corn starch (WC), common corn starch (CC), and 50% and 70% amylose corn starches (AMC)). Corn starches were surface gelatinized by 13 M LiCl at room temperature to different extents (approximately 10%, 20%, 30%, and 40%). The gelatinized surface starch and remaining granules were characterized for amylose content, amylopectin chain-length distribution, thermal properties, swelling power (SP), and water solubility index (WSI). Except for the outmost 10% layer, the amylose content in CC increased slightly with increasing surface removal. In contrast, amylose was more concentrated at the periphery than at the core for 50% and 70% AMC. The proportion of amylopectin A chains generally decreased while that of B1 chains generally increased with increasing surface removal for all corn starches. The gelatinization enthalpy usually decreased, except for 70% AMC, whereas the retrogradation enthalpy relatively remained unchanged for CC but increased for WC, 50% and 70% AMC with increasing surface removal. The SP and WSI increased with increasing surface removal for all corn starches, with WC showing a significant increase in SP after the removal of the outmost 10% layer. The results of this study indicated that there were similarities and differences in the distribution of amylose and amylopectin chains along the radial location of corn starch granules with varying amylose contents. More amylose-lipid complex and amylopectin long chains were present at the periphery than at the core for amylose-containing corn starches.     

Effect of aniline coupling on kinetic and thermodynamic properties of Fusarium solani glucoamylase

Purified glucoamylase (GA) from Fusarium solani was chemically modified by cross-linking with aniline hydrochloride in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) for 1 [aniline-coupled glucoamylase-1 (ACG-1)], 7 (ACG-7), and 13 min (ACG-13). The aniline coupling of GA had a profound enhancing effect on temperature, pH optima, and pK _a’s of active site residues. The specificity constants (K _cat/K _m) of native, ACG-1, ACG-7, and ACG-13 were 136, 244, 262, and 208 at 55°C for starch, respectively. The enthalpy of activation (ΔH*) and free energy of activation (ΔG*) for soluble starch hydrolysis were lower for the chemically modified forms compared to native GA. Proteolysis of ACGs by α-chymotrypsin and subtilisin resulted in activation.     

Physicochemical,Thermal, Pasting and Microstructure Properties of Hydroxypropylated Jackfruit Seed Starch Prepared by Etherification with Propylene Oxide

The jackfruit seed starch was subjected to the hydroxypropylation process by using various volumes of propylene oxide in the range of 5–50 ml (HP₅-HP₅₀). It was found that the molecular substitution (MS) increased with the volume of propylene oxide used. The shape of the native starch appeared irregular, round to bell or semi-oval with an average size of 7.66 μm. The modified granules were still intact in granular form without fragmentation. However, the presence of traces of degradation was observed on some starch granules, particularly the treatment of HP₄₀-HP₅₀. No marked differences in the XRD pattern were observed between the native starch and the hydroxypropyl derivatives. However, a decrease in the degree of crystallinity was found as the volume of propylene oxide in the reaction mixture increased. The swelling power and solubility of the hydroxypropylated starches were higher than in the native starch. Progressive increases in swelling power and solubility were observed as the MS increased among the hydroxypropylated starches. The pasting properties revealed that decreases in the pasting temperature, final viscosity and setback was found with increasing MS while the breakdown increased. Hydroxypropylated starches had lower gelatinization parameters (To, Tp, Tc and ?H) compared with the native starch. During refrigerated storage of the starch gel, the native starch showed syneresis from the first cycle while no syneresis was found during seven cycles for the treatment of HP₁₀-HP₅₀.     

Alteration of the properties of Aspergillus sp. K-27 glucoamylase on limited proteolysis with subtilisin

An active derivative (mol. wt. 48,000) of Aspergillus sp. K-27 glucoamylase (mol. wt. 76,000) was obtained by limited proteolysis with subtilisin. The amino acid sequences of native and modified enzymes at the N-termini were Ala-Gly-Gly-Thr-Leu-Asp and Ala-Val-Leu, respectively. The proteolysis greatly decreased the affinity of the enzyme for amylopectin and glycogen, but not for oligosaccharides. It also reduced the ability of the enzyme to degrade raw starch, abolished the ability of the enzyme to adsorb onto starch granules, and eliminated the synergistic action of the enzyme in the hydrolysis of starch granules with alpha-amylase. These findings imply that the enzyme has a specific affinity site for polysaccharide substrates besides the catalytic site, i.e., a starch-binding site, and that the former is removed by proteolysis. The extent of the reduction in the activity for raw starches caused by the modification varied with the starch source, as the modified enzyme digested raw potato starch better than either raw corn or sweet potato starches. A new method for evaluation of the raw starch-digesting activity of glucoamylase is described.