Capillary electrophoresis of complex natural polysaccharides

Abstract

Complex natural polysaccharides, glycosaminoglycans (GAGs), are a class of ubiquitous macromolecules that exhibit a wide range of biological functions and participate and regulate multiple cellular events and (patho)physiological processes. They are generally present either as free chains (hyaluronic acid and bacterial acidic polysaccharides) or as side chains of proteoglycans (PGs; chondroitin/dermatan sulfate, heparin/heparan sulfate, and keratan sulfate) and are most often found in cell membranes and in the extracellular matrix. The recent emergence of modern analytical tools for their study has produced a virtual explosion in the field of glycomics. CE, due to its high resolving power and sensitivity, has been useful in the analysis of intact GAGs and GAG-derived oligosaccharides and disaccharides affording concentration and structural characterization data essential for understanding the biological functions of GAGs. In this review, novel off-line and online CE-MS and MS/MS methods for screening of GAG-derived oligosaccharides and disaccharides will be discussed.

Novel and highly sensitive mixed‐polymeric electrokinetic chromatography system for determination of contaminants and impurities of heparin samples

“…In this regard, Volpi et al have presented CE-MS and MS-MS analytical methods for the screening of polysaccharidesderivated oligosaccharides and disaccharides as well as the analysis of low molecular weight heparins using 20 mM sodium phosphate buffer pH 3.48 with the addition of copper (II) sulfate. As a result of the addition of copper, the Hep peak was detected at 240 nm [3].…”

“…This is due to the fact that polysaccharides do not possess strong chromophores on their structures and have low extinction coefficients, making them difficult to measure by direct UV detection (3).…”

“…1). Hep has been used clinically as an anticoagulant and antithrombotic agent for many years [1][2][3].…”

Capillary Electrophoresis

“…Furthermore, the methodology used for the initial characterization of this contaminant (i.e., multiple orthogonal techniques, including multidimensional NMR [8]) is complex, expensive, and not useful for quality assurance in quality control laboratories because it is unable to process many samples in a short time and to give quantitative results with low coefficient of variation values. In contrast, capillary electrophoresis (CE) has been applied in the analysis of intact GAGs and GAG-derived oligosaccharides and disaccharides, such as monosaccharides, affording concentration and structural characterization data due to its high resolving power and sensitivity [12]. In this article, a validated CE method has been developed for the quantitative determination of OSCS in Hep preparations after degradation of the polysaccharides to produce hexosamine units (i.e., GlcN from Hep and GalN from OSCS), their derivatization with anthranilic acid (AA, 2-aminobenzoic acid), and separation by means of CE at 214 nm.…”

Proteoglycomics: Recent Progress and Future Challenges

“…In systematic studies such as proteoglycomics, disaccharide analysis is the first step in assessing changes in the GAG compositional profile. Disaccharide composition can be determined using a combination of on-line separation methods such as capillary electrophoresis (Amon et al, 2008;Volpi et al, 2008;Zaia, 2009), ion-pairing RP liquid chromatography (LC) (Doneanu et al, 2009;Korir et al, 2008;Zhang et al, 2009b), hydrophilic interaction LC (HILIC) (Zaia, 2009) in conjunction with MS or UV absorbance detection. Introducing a fluorophore by chemical derivatization of disaccharides greatly enhances their detection sensitivity (Deakin and Lyon, 2008;Hitchcock et al, 2008a;Skidmore et al, 2009) and is a basis for such detection methods as laser-induced fluorescence (Hitchcock et al, 2008b;Korir and Larive, 2009;Skidmore et al, 2009) and fluorophore-assisted carbohydrate electrophoresis (Volpi and Maccari, 2006).…”