Commercial Amylose Research Articles

Using Rice Flour for Purification of Maltose Binding Fusion Proteins Expressed in Escherichia coli

Background: One disadvantage of expressing heterologous proteins in Escherichia coli is that the proteins are frequently expressed as insoluble inclusion bodies. To avoid this problem, heterologous proteins are typically expressed as a fusion protein. Maltose binding protein (MBP) is one of the widely used partners for production of recombinant fusion proteins in E. coli. MBP is among the most effective solubility enhancers. In addition, MBP can be used as an affinity tag for purification of recombinant proteins on a column of amylose resin. Objectives: The purpose of this study was to investigate the use of rice flour, a natural source of amylose, for purification of MBP fusion proteins. Materials and Methods: MBP and a fusion protein of MBP and avian influenza virus nucleoprotein (MBP-NP) were expressed in E. coli and subjected to purification by rice flour and a commercial amylose resin. The purified proteins were analyzed by SDS-PAGE. Results: The results indicated that MBP and MBP-NP, both were successfully purified by rice flour. Conclusions: Rice flour can be used for purification of MBP fusion proteins. Although the efficiency of purification by rice flour is less than amylose resin, however, the yield is sufficient to obtain a quantity of protein required for research purposes.

Gel formation in mixtures of high amylopectin potato starch and potato starch

The effects of starch granules on the rheological behaviour of gels of native potato and high amylopectin potato (HAPP) starches have been studied with small deformation oscillatory rheometry. The influence of granule remnants on the rheological properties of samples treated at 90 °C was evident when compared with samples treated at 140 °C, where no granule remnants were found. The presence of amylose in native potato starch gave to stronger network formation since potato starch gave higher moduli values than HAPP, after both 90 and 140 °C treatments. In addition, amylose may have strengthened the network of HAPP because higher moduli values were obtained when native potato starch was added to the system. The moduli values of the mixtures also increased with increasing polysaccharide concentration in the system, which is due to an increment in the polysaccharide chain contacts and entanglements. Finally, it was found that a mixture of commercial amylose from potato starch and HAPP resulted in lower values of G′ compared to native potato starch. This indicates that the source of amylose is important for the properties in a blend with native amylopectin.

Gel formation in mixtures of high amylopectin potato starch and potato starch

The effects of starch granules on the rheological behaviour of gels of native potato and high amylopectin potato (HAPP) starches have been studied with small deformation oscillatory rheometry. The influence of granule remnants on the rheological properties of samples treated at 90 °C was evident when compared with samples treated at 140 °C, where no granule remnants were found. The presence of amylose in native potato starch gave to stronger network formation since potato starch gave higher moduli values than HAPP, after both 90 and 140 °C treatments. In addition, amylose may have strengthened the network of HAPP because higher moduli values were obtained when native potato starch was added to the system. The moduli values of the mixtures also increased with increasing polysaccharide concentration in the system, which is due to an increment in the polysaccharide chain contacts and entanglements. Finally, it was found that a mixture of commercial amylose from potato starch and HAPP resulted in lower values of G ′ compared to native potato starch. This indicates that the source of amylose is important for the properties in a blend with native amylopectin.

リゾホスファチジルコリンと複合体を形成するアミロース鎖長の解析

The formation of amylose complexes with eight lysophosphatidylcholines; 1-capryl lysophosphatidylcholine (LPC-C 10 : 0), 1-lauroyl LPC (LPC-C 12 : 0), 1-myristoyl LPC (LPC-C 14 : 0), 1-palmitoyl LPC (LPC-C 16 : 0), 1-stearoyl LPC (LPC-C 18 : 0), 1-oleoyl LPC (LPC-C 18 : 1), 1-linoleoyl LPC (LPC-18 : 2) and 1-arachidoyl LPC (LPC-C 20 : 0), was conducted in aqueous solution using a commercial amylose with an average degree of polymerization (DP) of 18. The chain distributions of amylose precipitated by forming a complex in aqueous solution (2°C) were analyzed by high-performance anion exchange chromatography with a pulsed amperometric detector (HPAEC-PAD). The smallest DPs of amylose forming complexes with LPC-C 14 : 0, LPC-C 16 : 0, LPC-C 18 : 0, LPC-C 18 : 1, LPC-18 : 2 and LPC-C 20 : 0 were DP 27, DP 29, DP 29, DP 31, DP 31 and DP 35, respectively. The principal amylose chains found in complexes with LPC-C 14 : 0, LPC-C 16 : 0, LPC-C 18 : 0, LPC-C 18 : 1, LPC-18 : 2 and LPC-C 20 : 0 were DP 34-36, DP 32-34, DP 33-35, DP 34-36, DP 36-38 and DP 36-38, respectively.

Characteristics of Amylose Forming Complexes with Glycerol Monooleate and Monostearate.

The formation of amylose complexes with glycerol monooleate (MG-C18:1) or glycerol monostearate(MG-C18:0) was conducted in aqueous solution using a commercial amylose with an average degree of polymerization (DP) of 18. The chain distributions of amylose precipitated by forming a complex in aqueous solution (20°C) were analyzed by high-performance anion exchange chromatography with a pulsed amperometric detector (HPAEC-PAD). The critical DPs of amylose forming complexes with MG-C18:1 and MG-C18:0 were DP 31 and 29, respectively. The principal chains of amylose forming complexes with MG-C18:1 and MGC18: 0 were DP 35-36 and DP 33-34, respectively.

Viscometric Properties of Solutions of Amylose and Amylopectin in Aqueous Potassium Thiocyanate

Solutions of commercial potato amylose (1.65%, w/v) in dimethyl sulfoxide (DMSO) /water mixtures formed gels on standing when the water content was equal to or greater than 60% (w/v). These gels did not form when the water was replaced with aqueous potassium thiocyanate solution (KSCN, 2 mol/L). Commercial amylose did not dissolve completely in aqueous KSCN, unlike commercial potato and corn amylopectins, which formed more stable solutions than those in water alone. Solutions of amylopectin in mixtures of DMSO and aqueous KSCN showed a maximum relative viscosity (flow viscometer) at 80% (v/v) DMSO, like those in DMSO/water. Solutions of amylopectin in 25% (v/v) DMSO/75% (v/v) aqueous KSCN (2 mol/L) could be used for estimation of molecular weight by flow viscometry using the Mark-Houwink-Sakurada Equation. Sigma corn amylopectin gave a weight average molecular weight of 12.1 x 106, lower than that obtained using DMSO alone, and likely to be due to a lower degree of hydrogen bonding in the presence of KSCN, consistent with its chaotropic effect. This effect was also reinforced by the observation that formation of the amylose/iodine complex is inhibited by KSCN at concentrations as low as 0.2 mol/L.

Isolation and Characterisation of Single Chain Amylose

Abstract Five amylose preparations of different origins (cassava, potato, smooth seeded pea, wheat and maize), obtained from native starch granules by thymol complexation, were ultracentrifuged as the final decontamination step to remove a high molecular weight population contaminating the amylose solutions. The efficiency of this ultracentrifugation procedure was assessed by dynamic light scattering (DLS): decreases in apparent hydrodynamic radii, R? H , from 46·1–72·6 nm before ultracentrifugation to 16·1–29·3 nm after were observed. Amylose solutions were then characterised by size exclusion chromatography coupled on-line to multi-angle laser light scattering (SEC–MALLS). Under these conditions M? w of cassava, potato, smooth seeded pea, wheat and maize amyloses were, respectively, 1·05×10 6 , 7×10 5 , 6·2×10 5 , 5·1×10 5 and 3·4×10 5 g/mol. Using a specific optimisation algorithm, experimental molecular weight distributions (MWD) were fitted by two mathematical models of ‘Most Probable’ distribution and ‘log-normal’ distribution. The best fit was obtained for the second model, but fitted M? w were higher than experimental M? w . When a ‘Most Probable’ model was used, the fitted M? w were consistent with experimental M? w but with a lower quality of fit. Exponents c in the power law R G = KM c were between 0·6 and 0·7, indicating an extended linear random coil in the range of MW analysed (3×10 5 –9×10 6 g/mol). This procedure was also applied to the characterisation of different commercial amylose products.

Nägeli amylodextrin and its relationship to starch granule structure. I. Preparation and properties of amylodextrins from various starch types

AbstractNägeli‐type amylodextrins were prepared from various starch types: native starch granules from potato, waxy maize, wrinkled pea, and high‐amylose maize, and from a commercial amylose (“Superlose”). The granular starches and retrograded amylose were treated with 15% sulfuric acid at 22°–25° for 3 months, the undissolved residues were washed free of acid and air dried. X‐ray diffraction patterns showed that amylodextrin from waxy maize starch (A‐type) and potato starch (B‐type) retain the same diffraction type as that of the parent starch. On conversion of a starch to an amylodextrin, the sharpness and intensity of the diffraction patterns are either retained (waxy maize starch), improved (slightly with potato, significantly with high‐amylose maize, and very markedly with wrinkled pea) or developed (amylose). The results indicate that the crystalline regions of a native starch granule, retrograded amylose, or amylodextrin are exceedingly resistant to acidic hydrolysis at room temperature. In contrast to the parent starches, native amylodextrins stain little if at all with dilute iodine solution. The iodine stains of dissolved amylodextrins were red (waxy maize), red‐purple (potato), or purple (amylose, high‐amylose maize, and wrinkled pea).

The interaction of polysaccharides with iodine : Part I. Investigation of the general nature of the reaction

A comparison of the interaction of a commercially available xylan, a d-galactose-deficient galactoglucomannan from Englemann spruce holocellulose, a highly branched “amyloid” from tamarind seed, and a commercial amylose preparation from potato starch, in concentrated aqueous calcium chloride solution showed that all four of these polymers reacted with iodine-potassium iodide solution to give a blue product that was soluble at low concentrations of reagents. In agreement with data in the literature, other highly branched polysaccharides such as cherry gum and d-galactose-rich galactoglucomannans did not reach with iodine under these conditions. Qualitative tests showed that iodine and calcium ion, as well as polysaccharide, were present in the isolated complexes, and spectrophotometric measurements showed the dependence of complex formation on the concentration of iodine and polysaccharide as well as on the time and temperature of reaction. Although the polysaccharides reacted to give a dark blue, starch-like coloration with iodine, potentiometric titration showed that only the xylan was complexed with iodine in a manner similar to that of amylose, while galactoglucomannan and “amyloid” bound iodine in a looser fashion typical of poly(vinyl alcohol)-iodine complexes. The iodine content of the complexes from xylan, “amyloid”, and galactoglucomannan, unlike the iodine content of the amylose complex, was found to vary with the concentration of the reactants, indicating the absence of a preferred stoichiometry.

Rotation of Reducing Power, as Measured by Fehling's Solution to the Rotatory Power of Commercial Amylose (Glucose and Grape Sugar)

Rotation of Reducing Power, as Measured by Fehling's Solution to the Rotatory Power of Commercial Amylose (Glucose and Grape Sugar)