title: New electrophoretic and chromatographic techniques for analysis of heparin and heparan sulfate.
abstract: Heparin (HE) and heparan sulfated glycosaminoglycans are well-known mediators of tissue development, maintenance and functions; the activities of these polysaccharides are depending mainly on their sulfate substitutions. The HE structure is also a very important feature in antithrombotic drug development, since the antithrombin binding site is composed by sequences of a specific sulfation pattern. The analysis of disaccharide composition is then a fundamental point of all the studies regarding HE/heparan sulfate glycosaminoglycan (and thereby proteoglycan) functions. The present work describes two analytical methods to quantify the disaccharides constituting HE and heparan sulfate chains. The use of PAGE of fluorophore-labeled saccharides and HPLC coupled with a fluorescence detector allowed in one run the identification of 90-95% of HE disaccharides and 74-100% of rat kidney purified heparan sulfate. Moreover, the protocol here reported avoid the N-sulfation disaccharides degradation, which may affect N-sulfated/N-acetylated disaccharides ratio evaluation. These methods could be also very important in clinical treatments since they are useful for monitoring the availability kinetics of antithrombotic drugs, such as low-molecular-weight HEs.
authors.affiliation: Department of Experimental and Clinical Biomedical Sciences, University of Insubria, Varese, Italy.
authors.forename: Manuela
authors.initials: M
authors.lastname: Viola
authors.forename: Davide
authors.initials: D
authors.lastname: Vigetti
authors.forename: Evgenia
authors.initials: E
authors.lastname: Karousou
authors.forename: Barbara
authors.initials: B
authors.lastname: Bartolini
authors.forename: Anna
authors.initials: A
authors.lastname: Genasetti
authors.forename: Manuela
authors.initials: M
authors.lastname: Rizzi
authors.forename: Moira
authors.initials: M
authors.lastname: Clerici
authors.forename: Francesco
authors.initials: F
authors.lastname: Pallotti
authors.forename: Giancarlo
authors.initials: G
authors.lastname: De Luca
authors.forename: Alberto
authors.initials: A
authors.lastname: Passi
mesh_terms: D000107:Acetylation; D000609:Aminoacridines / Q000032:analysis; D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005456:Fluorescent Dyes / Q000032:analysis; D006025:Glycosaminoglycans / Q000032:analysis; D006493:Heparin / Q000032:analysis* / Q000737:chemistry; D006497:Heparan Sulfate / Q000032:analysis* / Q000737:chemistry; D051381:Rats; D012680:Sensitivity and Specificity; D013552:Swine
chemical_list: D000609:Aminoacridines; D005456:Fluorescent Dyes; D006025:Glycosaminoglycans; D006493:Heparin; D006497:Heparan Sulfate; C013786:A73025; C072515:2-aminoacridone