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Asymmetrical flow field flow fractionation (AF4) has proven to be a very powerful and quantitative method for the determination of the macromolecular structure of high molar mass branched biopolymers, when coupled with multi-angle laser light scattering (MALLS). This work describes a detailed investigation of the macromolecular structure of native glycogens and hyperbranched α-glucans (HBPs), with average molar mass ranging from 2 × 10 to 4.3 × 10 g mol, which are not well fractionated by means of classical size-exclusion chromatography. HBPs were enzymatically produced from sucrose by the tandem action of an amylosucrase and a branching enzyme mimicking in vitro the elongation and branching steps involved in glycogen biosynthesis. Size and molar mass distributions were studied by AF4, coupled with online quasi-elastic light scattering (QELS) and transmission electron microscopy. AF4-MALLS-QELS has shown a remarkable agreement between hydrodynamic radii obtained by online QELS and by AF4 theory in normal mode with constant cross flow. Molar mass, size, and dispersity were shown to significantly increase with initial sucrose concentration, and to decrease when the branching enzyme activity increases. Several populations with different size range were observed: the amount of small size molecules decreasing with increasing sucrose concentration. The spherical and dense global conformation thus highlighted was partly similar to native glycogens. A more detailed study of HBPs synthesized from low and high initial sucrose concentrations was performed using complementary enzymatic hydrolysis of external chains and chromatography. It emphasized a more homogeneous branching pattern than native glycogens with a denser core and shorter external chains. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Starch consists of a mixture of two α-glucans built mainly upon α-(1,4) linkages: amylose, an essentially linear polymer, and amylopectin, a branched polymer containing 5-6% of α-(1,6) linkages. The aim of the present work was to analyze the structural properties of native starches displaying different amylose-to-amylopectin ratios and arising from different botanical sources, using asymmetrical flow field flow fractionation (A4F) and a combination of hydrodynamic and size-exclusion chromatography (HDC-SEC) coupled with multiangle laser light scattering, online quasi-elastic light scattering, and differential refractive index techniques. The procedure, based upon dimethyl sulfoxide pretreatment and then solubilization in water, generates a representative injected sample without altering the initial degree of polymerization. The amylopectin weight-average molar masses and radii of gyration were around 1.0 × 10-4.8 × 10 g mol and 110-267 nm, respectively. For each starch sample, the hydrodynamic radius ( R) distributions and the molar mass distributions obtained from the two fractionation systems coupled with light scattering techniques were analyzed. The size determination scales were extended by means of R calibration curves. HDC-SEC and A4F data could be matched. However, A4F enabled a better separation of amylopectins and therefore an enhanced structural characterization of the starches. The two advantages of this experimental approach are (1) it can directly obtain distributions as a function of both molar mass and size, while taking account of sample heterogeneity, and (2) it is possible to compare the results obtained using the different techniques through the direct application of R distributions. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]