Ionic strength effect on molecular structure of hyaluronic acid investigated by flow field-flow fractionation and multiangle light scattering

Abstract

This study describes the effect of ionic strength on the molecular structure of hyaluronic acid (HA) in an aqueous solution using flow field-flow fractionation and multiangle light scattering (FlFFF-MALS). Sodium salts of HA (NaHA) raw materials (∼2 × 10(6) Da) dispersed in different concentrations of NaCl prepared by repeated dilution/ultrafiltration procedures were examined in order to study conformational changes in terms of the relationship between the radius of gyration and molecular weight (MW) and molecular weight distribution (MWD) of NaHA in solution. This was achieved by varying the ionic strength of the carrier solution used in a frit-inlet asymmetrical FlFFF (FIAF4) channel. Experiments showed that the average MW of NaHA increased as the ionic strength of the NaHA solution decreased due to enhanced entanglement or aggregation of HA molecules. Relatively large molecules (greater than ∼5 MDa) did not show a large increase in RMS radius value as the NaCl concentration decreased. Conversely, smaller species showed larger changes, suggesting molecular expansion at lower ionic strengths. When the ionic strength of the FlFFF carrier solution was decreased, the HA species in a salt-rich solution (0.2 M NaCl) underwent rapid molecular aggregation during FlFFF separation. However, when salt-depleted HA samples (I = 4.66∼0.38 mM) were analyzed with FFF carrier solutions of a high ionic strength, the changes in both molecular structure and size were somewhat reversible, although there was a delay in correction of the molecular structure.

Effect of electrostatic interactions on the ultrafiltration behavior of charged bacterial capsular polysaccharides

“…The solutions were highly shear‐thinning and showed significant viscoelastic behavior. Kim et al used field flow fractionation and multi‐angle light scattering to investigate the structure of hyaluronic acid, with the results showing both aggregation and charge‐induced molecular expansion depending upon the solution ionic strength and the molecular weight of the polysaccharide. Choppe et al showed that the relaxation time for xanthan gum increased by more than two orders of magnitude when 100 mM NaCl was added at low temperatures.…”

Correlation Analysis of Chitosan Physicochemical Parameters Determined by Different Methods

“…Problems of MW determination are not unique for chitosan. Contrary to DNA and proteins, which have stable conformation, polysaccharides are prone to change it in different conditions (salt, temperature, pressure) and can easily form complexes [46][47][48][49][50]. Our results demonstrate that MW may not be considered as the best characteristic of chitosan.…”

Factors determining self-assembly of hyaluronan

“…for fractionation of molecular mixtures, which allows avoiding many problems in polymer analysis related to the stationary phase such as unspecific adsorption and extensive shear forces applied on large macromolecules. A FFF sub-technique, asymmetric flow field-flow fractionation (AF4) allows a fractionation of complex polymer systems according to molecular size by application of longitudinal and crossoriented flows in the channel (Kim et al, 2015;Malik & Pasch, 2016;Shin, Hwang, Cho, & Moon, 2007). The hyaluronan used in the study was characterized first using AF4 connected to MALS detectors (Wyatt, 1997) in order to obtain molecular masses, polydispersity and shape factors of the polysaccharide chains in solutions (Table 1).…”

“…HU networks properties have already been analyzed by various techniques, e.g. rotary shadowing electron microscopy (Scott et al, 1991), X-ray crystallography (Hargittai & Hargittai, 2008), NMR spectroscopy (Almond, DeAngelis, & Blundell, 2006;Cowman, Hittner, & Feder-Davis, 1996;Nestor & Sandström, 2017;Scott & Heatley, 1999), light scattering (Kim, Woo, Park, Hwang, & Moon, 2015), tensiometry (Musilová, Kašpárková, Mráček, Minařík, & Minařík, 2019), chromatography (Shanmuga Doss, Bhatt, & Jayaraman, 2017), molecular dynamics (Cowman & Matsuoka, 2005;Hargittai & Hargittai, 2008;Kutálková, Hrnčiřík, Witasek, & Ingr, 2020) and atomic force microscopy (AFM) (Cowman, Li, & Balazs, 1998, 2005Giannotti, Rinaudo, & Vancso, 2007;Gunning, Morris, Al-Assaf, & Phillips, 1996;Jacoboni, Valdrè, Mori, Quaglino, & Pasquali-Ronchetti, 1999;Spagnoli et al, 2005;Zellermann et al, 2013). It has been proven, that despite the effect of the surface and adsorbed water described by , AFM can be used to study the aggregation and networking ability of HU with respect to the process conditions of solution preparation.…”