C.T. Greenwood

The interaction of linear, amylose oligomers with iodine

A series of linear, amylose oligomers has been synthesized enzymically to yield samples having the number-average degree of polymerisation, DPn¯, in the range 22 to 134. The interaction of these materials with iodine has been studied. The wavelength of maximal absorption of the iodine complex, λmax, has been found to be related to DPn¯ by the Langmuir isotherm: 1/λmax⁡=1.558×10−3+102.5×10−4(1/DPn¯). Differential, potentiometric titration of iodine has shown that the iodine-binding capacity of the linear oligomers is a function of both DPn¯ and temperatures. The enthalpy of the iodine uptake has been measured, and found to be a function of DPn¯ for values of DPn¯<200.

Studies on starches of high amylose-content: Part X. An improved method for the fractionation of maize and amylomaize starches by complex formation from aqueous dispersion after pretreatment with methyl sulphoxide

Abstract A method has been developed of separating maize starches of different genotypes quantitatively into their component material. Pretreatment of the granular starches with methyl sulphoxide is followed by the formation of a stable, aqueous methyl sulphoxide dispersion from which the amylose component can be rapidly precipitated as an easily centrifugeable complex with butan-1-ol. The residual starch material has been subfractionated by the formation of the iodine complex at low temperatures. Properties of the different starch fractions from this fractionation scheme have been examined, and the results are briefly considered with regard to current theories of maize starches of high amylose-content.

The conformation of amylose in neutral, aqueous salt solution☆

Abstract Linear amylose has been subfractionated, and the hydrodynamic properties of the fractions examined. From measurements of the limiting-viscosity number, [ n ], in 0.33m potassium chloride, and of the weight-average molecular weight, M w , by light scattering on the acetate, the relation was found. The significance of the exponent in this relation and others in the literature is discussed. Estimations of the “stiffness” of the amylose molecule in aqueous solution have been obtained for various model systems, i.e., the Kuhn statistical length, the Kratky-Porod persistence length, and Nagais theory for a mixture of helical and randomly coiled segments. The results of each individual theory are ambiguous. It is shown, on the basis of Nagais theory, that there is definite evidence for the absence of rigid helical segments in amylose in aqueous solution. The temperature coefficient of [ n ] for amylose in aqueous solution is also discussed.

The hydrodynamic behaviour of native amylose in good solvents

Abstract Viscosity measurements on 12 fractions of linear amylose have been made in three thermodynamically good solvents—formamide (F), methyl sulphoxide (Me2SO), and aqueous potassium hydroxide (KOH). The following Mark-Houwink relations were obtained: [ η ] F = 3.05 × 10 − 2 M ¯ ω 0.62 , [ η ] M e 2 S O = 1.51 × 10 − 2 M ¯ ω 0.70 , and [ η ] K O H = 8.36 × 10 − 3 M ¯ ω 0.77 . The Kamide-Inamoto and Stockmayer-Fixman-Burchard theories show that the amylose molecules in these solvents are extended as a result of solute-solvent interaction and there is little evidence of any free-draining characteristics. The graph of [η]/M1/2 againstM1/2 for all three good solvents is linear with a common intercept. Incidently, this intercept is the same as that for an aqueous theta-solvent, showing that the basic skeletal structure of amylose is the same in each of these solvents. The conformation of amylose in these three good solvents is regarded as being that of a random coil with negligible helical character.

Studies on starches of high amylose-content. : Part VIII. The effect of low temperature on the interaction of amylomaize starch with iodine: a unique characterization

Abstract The iodine-binding characteristics of amylomaize starch are affected dramatically by a lowering of temperature. At 2°, a normal-shaped, potentiometric, iodine-titration curve is obtained, from which, in contrast to results at 20°, estimation of the iodine affinity can be readily made. These conditions provide a unique method of characterization of this type of starch. Amylomaize starches having a variety of reported amylose-contents have been examined by this technique. The iodine-affinity values indicated that the starches contain some 15–18% more linear-material than expected. The iodine affinity at 2° of normal amylose and amylopectin is not appreciably different from that at 20°, but very large changes occur in the iodine-binding characteristics of samples of 36-unit amylopectin and degraded amylose at this low temperature. It is suggested that these changes account for the abnormal behaviour of the whole amylomaize starch.

The conformation of amylose in alkaline salt solution

Abstract The conformation of amylose in various solvents is discussed. It is shown that the changes in molecular volume of the polysaccharide (measured by viscosity) as potassium chloride is added to a solution of amylose at pH 12 are similar to those obtained on adding butan-1-ol to the solution. The viscosity number in both cases decreases to values less than that observed for amylose in water, in which Flory theta-conditions are approximated. The minimum value of the viscosity number, in fact, is identical to that observed on the addition of butan-1-ol and iodine to neutral aqueous solution of amylose—conditions known to result in a helical complex. It is concluded that amylose undergoes a coil-to-helix transition as potassium chloride is dadde to solutions of the polysaccharide at pH 12.

A novel method of determining the number-average degree of polymerization of linear maltodextrins and amylose

Abstract An enzymic method for determining the number-average degree of polymerization ( D P ¯ n ) of maltodextrins and amylose is described. In this technique, the D -glucans are treated with β-amylase under conditions in which enzymic degradation is complete, i.e., molecules having an even number of residues form maltose, and molecules having an odd number of residues yield maltose and one D -glucose unit. The amount of D -glucose is estimated by using glucose oxidase. The total concentration of the D -glucan itself is measured by use of amyloglucosidase. Hence, on the basis that the sample contains equal numbers of molecules having odd and even numbers of D -glucose residues, D P ¯ n can be calculated. Experimental conditions for this assay are discussed, and the accuracy of the technique has been confirmed by using (a) pure maltodextrins, and (b) synthetic mixtures of two degraded amyloses of D P ¯ n ca. 400 and 8. The method has been found to be applicable to linear amylose; D P ¯ n values greater than 1000 can be obtained to within 5%.

Viscosity and sedimentation studies on amylose in aqueous solution—further evidence for non-helical character☆

Abstract Viscosity and sedimentation measurements have been carried out on subfractions of native amylose dissolved in a thermodynamically ideal solvent (0·33 M KCl) and a good solvent (0·2 M KOH). The relations: [ƞ] KC1 = 1·12 x 10 -1 M W 0.50 ; [ƞ] KOH = 6·92 x 10 -3 M W 0.78 ; ( S 0 ) KC1 = 1·11 x 10 -15 M W 0.50 ; ( S 0 ) SOH = 2·73 x 10 -15 M W 0.41 ; were obtained. The divergence of the exponents from a value of 0·50 for the measurements in KOH is shown to be due to expansion of the molecule through solvent-polymer interaction rather than to partial free-draining character. The unperturbed dimensions of amylose were found to be the same in both solvents, on the basis of the Burchard-Stockmayer-Fixman and the Cowie-Bywater treatments. Measures of ‘molecular stiffness’ were obtained (Kuhn-Kuhn and Hearst-Stockmayer theories) on the basis of different molecular models including that of the ‘interrupted helix’. The latter model appears to be incorrect: analysis of the hydrodynamic data shows that the amylose molecule in aqueous solution is best regarded as a fairly flexible coil.

Studies on starch-degrading enzymes : Part II the Z-enzyme from soya beans; purification and properties

Abstract A method for the isolation and purification of the Z-enzyme in soya beans is described. This procedure involves the formation of the glycogen-enzyme complex. A method for assay of activity is presented. The properties of the purified enzyme have been studies; in particular, the variation of activity and stability with temperature and pH has enabled the nature of the active sites in the enzyme to be investigated. Characterization of the activity of Z-enzyme at pH 3.6, and in the presence of a variety of reagents, has been achieved by viscometric techniques. Both linear and branched glucans are attacked by the purified enzyme. It is concluded that the properties of Z-enzyme are similar to those of other plant α-amylases.

Action pattern of broad bean α-amylase☆

Abstract A study was made of the mode of action of broad bean α-amylase on linear amylose. Viscometric techniques were used to show that the initial hydrolysis appears to be random. The products of the degradation of amylose at the achroic limit, and also of the amylolysis of individual small oligosaccharides, were investigated by paper chromatography. The results indicate that the attack is nonrandom in the later stages. Quantitative determinations of the proportions of maltodextrins produced after the achroic limit confirmed the nonrandom nature of the enzyme action. Yields of dextrins to be expected at various stages of the degradation were calculated for different theories of amylase action and compared with the experimentally determined values. A reaction scheme has been proposed to account for the observations.