Low‐angle laser light scattering–aqueous size exclusion chromatography of polysaccharides: Molecular weight distribution and polymer branching determination

Abstract

SynopsisA low-angle laser light scattering detector (LALLS) used with size exclusion chromatography (SEC/LALLS) has been applied for the determination of molecular weight, molecular weight distribution (MWD), and degree of branching of polysaccharides in 0.5N NaOH aqueous solution. Data from both detectors [differential refractive index (DRI) and LALLS] are used to calculate the absolute molecular weight at each point in a sample chromatogram. The correct average molecular weight and MWD can be obtained without calibration methods used in conventional SEC. As a consequence of this technique, Mark-Houwink coefficients can be predicted from a single broad-distribution, homopolymer without recourse to timeconsuming fractionation methods. Moreover, the hydrodynamic volume separation mechanism of SEC can be exploited with the SEC/LALLS method to gain information about polymer branching. In the studies described in this paper, SEC/LALLS has been employed to obtain data about the branching parameters g, and g , , , for samples of amylose, amylopectin, starch, and glycogen. For three homopolymers (amylose, amylopectin, and glycogen), branching frequency (as measured by chemical means), and the branching parameters (go and gM) are inversely related. This trend is consistent with theoretical predictions. For starch, a nonhomogeneous branching distribution is observed as a function of molecular weight.

Enzyme-Synthesized Highly Branched Maltodextrins Have Slow Glucose Generation at the Mucosal α-Glucosidase Level and Are Slowly Digestible In Vivo

“…The collected data from MALS and RI detectors were analyzed using a Berry plot for curve fitting with Astra software version V (Wyatt Tech. Corp., Santa Barbara, CA), and a dn/dc value of 0.146 mL/g was applied for M w calculation [32], [33].…”

Application of size-exclusion chromatography/low angle laser light scattering in fermentation processes

“…In both cases, samples of the fermentation broth were taken at 12 hour intervals and centrifuged to remove the bacterial cells (10,000 RPMs for 30 minutes), decanted, and analyzed by SEC/LALLS. The specific details of the SEC/LALLS apparatus are described elsewhere (Yu and Rollings, 1987).…”

“…The utility of SEC/LALLS for polysaccharide characterization (Yu and Rollings, 1987;Yu and Rollings, 1988;Cael et al, 1983) has been previously demonstrated. Size exclusion chromatography separates molecules based on their hydrodynamic size in dilute solution.…”

“…By coupling various detection devices down-stream from an SEC separation, additional information about the chromatographed fermentation broth components can be obtained. The general operation of this device is described elsewhere (Yu and Rollings, 1987). The applicability of this technique for studies of polysaccharide depolymerase production processes is described here.…”

Effects of solution conditions on characteristics and size exclusion chromatography of pneumococcal polysaccharides and conjugate vaccines

“…Size exclusion chromatography (SEC) has been used extensively for characterization of carbohydrate polymers, including capsular bacterial polysaccharides (and their conjugates) that are used as important vaccine products to ensure both the stability and effective immunogenicity (Gaborieau & Castignolles, 2011;Lee et al, 2009;Mori & Barth, 2013;Pato, Antonio de Pádua & da Silva Junior, 2006;von Hunolstein, Parisi & Bottaro, 2003;Yu & Rollings, 1987). For example, Lee et al (2009) used size exclusion chromatography for the purification and analysis of a meningococcal conjugated vaccine.…”