Heparin chromatography

Affinity chromatography is used in the preparation of more highly purified Factor IX concentrates (53—55) as well as in the preparation of products such as antithrombin III [9000-94-6] (56,57). Heparin [9005-49-6], a sulfated polysaccharide (58), is the ligand used most commonly in these appHcations because it possesses specific binding sites for a number of plasma proteins (59,60). 

Lipoprotein lipase (from bovine skimmed milk) [9004-02-8] [EC 3.1.1.34]. Purified by affinity chromatography on heparin-Sepharose [Shirai et al. Biochim Biophys Acta 665 504 1981]. 

Pituitary Growth Factor (from human pituitary giand) [336096-71-0]. Purified by heparin and copper affinity chromatography, followed by chromatography on carboxymethyl cellulose (Whatman 52). [Rowe et al. Biochemistry 25 6421 1986.]... 

Ruiz-Calero, Puignou, L., Galceran, M. T., and Diez, M, Coupling high-performance size exclusion and ion chromatography for the analysis of low-molecular-mass heparin, /. Chromatogr. A, 775, 91, 1997. 

A semi-interpenetrated network was obtained by bulk polymerization of 2-hydroxye-thyl methacrylate incorporated in DMF treated PET films by solvent-exchange technique, followed by treatment of films in e-lectrical discharges. Heparinization was accomplished by reacting glutaraldehyde with heparin and poly(2-hydroxyethyl methacrylate) present on the surface of modified polyester films. The immobilization of heparin was indirectly evidenced by chromatographying the silylated hydrolyza-tes of heparinized PET films and heparin, respectively. In vitro experiments demonstrated the enhanced thromboresistance of heparinized films. 

Although liquid chromatography (l.c.) under elevated pressure is now used essentially for determining the molecular weight of heparin,74,75 as an alternative to methods using soft gels,76 it can undoubtedly be used as well for rapid analysis of mixtures of glycosaminoglycans. 

From the foregoing discussion, it is apparent that heparin may be subdivided into an unlimited number of fractions when different separation approaches are applied to each fraction obtained by another procedure. More than a hundred fractions have been obtained by sequential use of affinity chromatography on antithrombin, precipitation with barium, and isoelectrofocusing.214 Although these fractions can scarcely be referred to as species, such an extensive fractionation stresses the concept of the heterogeneity of heparin, and the influence of minor differences in chemical constituents, or chain length, or both, on the physicochemical (and, conceivably, biological) properties of this polysaccharide. 

The clearing effect of heparin on blood is associated with release of the triglyceride-hydrolyzing enzyme lipoproteinlipase (LPL) from the surface of endothelial cells.458,459 In addition to a number of apparently equivalent lipases from different tissues,459 heparin also releases a hepatic lipase.460,461 As suggested by the results of affinity-chromatography studies, this release is probably associated with binding of the polysaccharide to the enzyme. 62,463 Because other polyanions,464 including the... 

A modified technique consisting of an electric field applied perpendicular to a flowing buffer solution and using chromatography paper as support(cross-electrophoresis) has been exploited to study the reaction of neomycin with heparin 9 184. Evidence for the formation of a neomycin-heparin complex was obtained by this means. 

A detailed examination of the affinity of SLPI for the heparinized capillary was next made using a stepwise elution (from 0.1 to 0.9 M NaCl) (Fig. 11). SLPI eluted from the capillary with 0.2 M NaCl. This agreed well with results obtained by traditional affinity chromatography on a heparin-Sepharose matrix. The ACE method has the unique advantages over traditional affinity chromatography in that it requires much smaller quantities of protein and afforded better separation profiles. 

X Wu, RJ Linhardt. Capillary affinity chromatography and affinity capillary electrophoresis of heparin-binding proteins. Electrophoresis 19 2650-2653, 1998. 

Degradation of heparin followed by affinity chromatography on immobilized AT III led to the identification of a pentasaccharide unit with high antithrombin affinity. This pentasaccharide 3 is characterized by a imique highly sulfated central glucosamine unit with a 3-0-sulfate group (Structures 2). 

By gel chromatography on Sephadex G-200, with an eluant consisting of 0.15 M sodium chloride mixed with 0.12 M sodium phosphate buffer solution, pH 7.4 (9 1, v/v), heparin of porcine mucosal origin has been shown to be highly polymolecular.127 No increase in blood anticoagulant activity with increasing molecular size, indicated by the results of an earlier study,128 was observed when the activity of fractions selected from different parts of the elution curve was determined. The anticoagulant potency of the sample studied was,... 

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